NUTRITIONAL AND GENETIC FACTORS CAUSING BENT-NOSE
IN THE RAT (RATTUS NORVEGICUS)

By
Walter E* Heston

A THESIS
presented to the Graduate School of Michigan State
College of Agriculture and Applied Science
in Partial Fulfillment of Requirements
for the Degree of
Doctor of Philosophy

Zoology Department
East Lansing, Michigan
1936

ProQuest Number: 10008330

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ACKNOWLEDGMENTS
/

The author wishes to express his appreciation to 8&1 who were
of assistance in this work*
Special gratitude is due Dr* H. R. Hunt of the Department of
Zoology for his helpful suggestions throughout the work and. especially
his guidance of the genetic experiments, also Dr* C« A* Hoppert of the
Chemistry Department under whose fine supervision the nutritional work
was done*

±05398

!EABLE OF CONTENTS
Page
X

Introduction...............................................

1

II

History of Bent-nose.......................................

6

III

Nutritional Factors Causing Bent-nose *••....................

8

1*

Introduction ........................................

8

2*

Eat ions U s e d .................................. .....

9

3*

Procedure......................................

11

H.

Presentation and Discussion ofD a t a

12

a.

Growth study.................................

12

b.

Mortality study

15

c.

Study of rations in relation to production

IV

...............................

of bent........................................

17

d.

Comparison of growth and incidence of bent-nose..**

21

e.

Bone-ash studies

23

f.

Results of subjecting animalsto a deficient

g.
5.

..............

.......... ...........

ration at 16 days of a g e ..........

yi

Results of inbreeding attempt...................

31

Summary of Nutritional Studies

.............

^0

Genetic Factors Causing Bent-nose.................

U2

I. Introduction.........................................

b2

2. Procedure for Studying the Genetic Factors Causing
Bent-nose in the Hat .................................
3* Presentation and Discussion of D a t a ....................
a*

Data obtained in nutritional studies

b*

Data obtained in genetic studies

U3
**5

♦• ^5
*...........

^7

4* Summary of Inheritance Studies........................

32

TABLES
Page
Table I

Average Weekly Weights of Groups of Animals Fed on
the Seven Different Hations «...........................

Table II

Mortality of Animals Fed on Ration II - High Calcium
Ration..........

Table III

*...... 16

Influence of the Seven Rations on the Frequency of
Bent-nose...............

Table IV

13

18

The Effect of Each Ration on the Frequency of Bent-nose
as Compared with its Effect on Growth R a t e ............... 22

Table V

Per Cent of Bone Ash in the Femurs of Animals Fed 20 Days
on Ration II - High Calcium Ration

Table VI

26

Per Cent of Bone Ash in the Femurs of Animals Fed *40 Days
on Ration II - High Calcium Ration

Table VII

....................

...................

27

Per Cent of Bone Ash in the Femurs of Animals Fed 60 Days
on Ration II - High Calcium Ration *....... .......... ♦.* 28

Table VIII Per Cent of Bone Ash in the Femurs of Animals Fed 20 Days
on Ration VII - High Phosphorus Ration
Table IX

................

29

Per Cent of Bone Ash in the Femurs of Animals Fed *40 Days
on Ration VII - High Phosphorus Ration................... 30

Table X

Ages at which Progeny of Bent x Bent Matings Died when put
on Ration II (High Calcium Ration) at Sixteen Days of Age
and Weaned at Twenty-one Days of Age

Table XI

........

32

Frequency of Bent-nosed Animals Among those of the First
Generation of Inbreeding

..........................

3^

TABLES

(Cont inued)
Page

Table XII

Comparison of Mean Weights of Bent-aosed Animals and
Rormal Animals of the Inbreeding Experiment

.............

Table XIII Wei^its of Hormal Animals in Inbreeding Experiment........
Weights of Bent-nosed Animals in Inbreeding Experiment
Table XIV

35
3&~3S
39

Frequencies of Bent-nose Among Animals from the
Various Crosses, All Reared on Ration VII

............. **9

ILLUSTRATIONS

Page
Figure

!• External Appearance of a Bent-nosed Animal .............

U

Figure

2* Skulls Showing the Bent-nosed Condition

5

Figure

3> Growth Curves.

.........

1*4-

INTRODUCTION
An organism Is the resultant of a multitude of processes, many
6f which are

initiated hy agencies within the fertilized egg, while others

are the consequence of external forces which impinge upon the individual
during development or later life*

The reciprocal relations between these

external and internal agents are usually complex and as a rule but little
understood*

Thus, ‘•inherited11 characters may be suppressed, stimulated, or

modified by “environmental•• forces*

The geneticist must always keep in

mind the multiple causations of traits; he must remember that light, temper­
ature, food, moisture, etc*, as well as the genes create the organism; in
short, he must never forget that he is a biologist*

Some inherited traits

are but little influenced by ordinary fluctuations in the environment,
while others are profoundly affected, as the following illustrations prove*
The development of the pattern of the Himalayan rabbit-^ is due
not only to the genetic constitution of the rabbit but also to the temperature to which the animal is subjected*

Bar-eye21

and vestigial wing2^+*29
*

are classic examples of genetic characters in Drosophila which are influenced
by temperature*

The character “abnormal abdomen" in Drosophila,

although

due to a dominant factor, is manifested only in flies which not only have
this factor but also have been reared on a moist culture medium*

Many other

characters could be cited in this connection, some involving environmental
factors other than temperature or moisture*
Bent-nose in the rat is another character which is affected not
only by genetic factors but by environmental agents as well*

The manifestation

of bent-nose depends on the proper genetic constitution in conjunction with

2-

an imbalance in the calcium-phosphorus ratio In the diet#

So essential

are both of these types of causative agents that neither could be analyzed
without due consideration of the other*
The name "bent-nose" was chosen because it described the character
simply and adequately*

(See figures 1 and 2)*

to the right or the left*

The rostrum was bent either

With the exception of the abnormalities common

to rachitic animals, such as the rachitic rosary, pigeon breast etc, no
other bone anomalies normally occurred in the bent-nosed animals*

Thus, it

can be stated that bent-nose is characterized by the bending of the rostrum
of the skull*
ik
Preliminary investigations
showed that the character did not
manifest itself until after birth*
was thirty-four days of age*

The youngest animal to show the trait

No animals have been known to develop the

character after they were 120 days old, and in most of the rats the anomaly
appeared between the age of sixty and seventy days*

The degree of flexure

was variable, ranging in a group of sixty-six bent-nosed skulls from U° to
as much as 35? with 17*5° as the mean grade of flexure.

The frequency of

the left bends was not significantly different from that of right bends,
and there was no statistically significant difference between the mean
angles to the left and right.

Of the sixty-six bent skulls, thirty-one

were bent to the left, and thirty-five to the right*

The mean angle of bend

to the left was 17*S? while that to the right was 17*3°*

difference

between these two means is 0*5°± 1.016 which is only 0*^*9 times its
probable error*

-3'

No description of such, a character in rats has been found in
the literature.

However* Green and apekete^® have described two mice which

showed a condition identical in external appearances to the bent-nose of
the rat.

The character might also be compared, to wtwisted beak11* in fowls?

reported be Hutti^ This is a sub-lethal character which appears to be
inherited as a simple recessive.

-H-

Figure 1
External appearance of a,
bent-nosed animal. Adult
animal with nose bent to
the right. (M.S. thesis)

Figure 2
Skulls showing the bent-nosed condition.
9 6 nomal. ? 7 » $ 9 aI*d 9 11 bent.
The absence of the zygomatic bone
is not an anomaly as in each case
it was lost while the skull was
being cleaned.
(M. S. thesis)

-6HISTOKY OP BENT-NOSE
Bent-nose was discovered during 1930* *n
laboratory at Michigan State College#

Elementary Zoology

Same of the animals that were being

used for mammalian dissection showed the anomaly*

These were rats from

the rodent colony of the Zoology Department at Michigan State College*
Immediate investigation of the different stocks in the colony
revealed that the character appeared only in the red-eyed-yellow strain
derived from Dr* W* E* Castle's stock, and that it was quite prevalent in
that strain which was a good indication that the character was inherited*
In 1931 and 1932 Mr* L* W* Wiren made some matings of bent x normal from
bent stock, and bent x bent, but the data he obtained were inadequate for
any definite conclusions*
During 1932» matings were made of the various types! bent x bent,
bent x normal,

x F^, and F^ x bent*

The stock was fed on the colony rat

rations consisting in general of a cooked diet of various grains with fruit
and meat scraps sometimes added*
cabbage or alfalfa*

Occasionally they were given green lettuce,

Two hundred and four animals were reared, of which 93’

or ^5*6 per cent, were bent*

Then on January 1, 1933>

r&ts in the

colony were put on the Purina Fox Chow ration as an economy measure*

One

hundred and ninety-one animals from the same types of matings were reared on
this diet, and not one had a bent nose* An inherited trait had disappeared!
This was ample proof that food wets a factor in the development of this trait*
Knowing that diet was a factor, it was desirable to discover what
nutritional agents favored the appearance of bent-nose, and to develop a
ration that could be used in studying the genetic factors involved*

If

genes for bent-nose could produce their effects only under a certain type

of diet, then that diet would have to he used throughout the genetic
experiments*

An appropriate research procedure was accordingly formulated*

The nutritional experiments were done under the supervision of Dr* C* A*
Eoppert of the Chemistry Department*

NUTRITIONAL FACTORS CAUSING BENT-NOSE
Introdactlon
To facilitate the production of a hone abnormality, one would
normally experiment with the two main hone—forming elements of the diet,
namely, calcium and phosphorus* Some of the early works of Steen^c^*^*
111,1^2,^4. _
35>36>37
Sherman,
and their colleagues have shown that an imbalanced
calcium-phosphorus ratio, in the absence of vitamin D, gives rise to rickets,
a condition often accompanied by various bone defects*

Such a rickets-

producing ration could be made according to several different formulae*
amount of calcium could be high with the phosphorus low —

The

the type of ration

usually employed; the amount of phosphorus could be high in relation to the
calcium; or both could be present in too small amounts*

Absence of vitamin

D is essential in any case since its presence tends to correct the imbalance
in the calcium-phosphorus ratio,

or in the case of a deficiency of both of

the elements in question, vitamin D permits a more efficient utilization of
these elements*
Some of the more recent works have shown that other factors may
affect calcification*

Sabel, Cohen, and Kramer^*’^

demonstrated that

strontium may cause rickets due to the injury it inflicts on the calcifying
mechanism*

H* V* Smith, M* C. Smith, and others32,3S>39, have shown that

fluorides also may be deleterious to calcification*
tend to be centered in the enamel of the teeth.
may cause a change in the calcification of teeth*

The effects of fluorides

Even a lack of vitamin A
The work of Smith and

-9-

Geiger, Staahbock, and Persons** reported a condition in the rat
known as "Lathyrism" in which calcification has been interfered with*

Quoting

from their paper: "Lathyrism was produced in both young and adult rats byfeeding diets which contained Lathyrus odoratus* the flowering sweet pea*
"Growth of the young animals was retarded by sweet peas when they
were fed 80$, 50$ an^> 25$ of the diet*

However, normal growth was obtained

for 20 weeks with 12*5$ and 5$ of sweet peas*

Other symptoms of lathyrism

noted were lameness, spinal curvature, sternal curvature, enlargement of the
costochrondral junctions, and malformation and abnormal red color of the
long bones*

Calcification was interfered with in young animals*n

More stress was laid on the calcium, phosphorus and vitamin D
constituents of the diet, in our study of bent-nose, than on the various other
elements Influencing calcification*
Rations Used*
Seven different rations were used in our experiments on the food
factors involved*

These included one well-balanced stock ration as a control

diet, and six deficient rations.

The ration which would produce the hipest

incidence of bending among progeny of bent x bent matings, and still not be
so deleterious that the animals died before they were 120 days of age, was
to be the ration later used in the breeding experiments*
the formulae for these different rations:
Ration I

- Stock ration

Corn meal
Oat meal
Milk powder
Oil meal
Alfalfa
Teast
NaCl

- JO parts
- JO parts
- JO parts
- 6 parts
- 5 parts
- 3 parts
- 1 part

28.57$)
28.57$)
28.57$)
5.71$)
&.7G$)

2.86$)

0.95$)

The following are

-10-

Bation II

- High, calcium ration

Corn meal
Oat meal
Wheat gluten
Ca CO
Ha Cl
Bation 1X1

- 38^
- 38^
- 20$
- 35^
- VL

- High calcium ration plus cod liver oil*

This ration was the same as Bation IX with 1 gram
of cod liver oil added to each 100 grams of
Bation II*
Bation IV

-

Yeast, high calcium ration

C o m meal
Oat meal
Wheat gluten
Ca CO,
Yeast^
Ha Cl
Bation Y

-

3#
3^
20J&
2#
Vjo
1#

- Irradiated yeast, high calcium ration

Corn meal
- 3*$
Oat meal
- 3^
Wheat gluten - 20^
Ca CO
2$
Irradiated yeast- 1$
Ha Cl
Vf>
Bation VI

- High phosphorus ration

C o m meal
Oat Meal
Wheat gluten
Oil meal
Casein
Yeast
Milk powder
Ha Cl
Bation VII

-

-

-

32$
10$
10%
5#
5#
%
¥

- High phosphorus ration

C o m meal
Ground wheat Wheat gluten Alfalfa
Ha Cl
-

U3$
hjjfi
10$
yfi
l£

-11-

Ration I, the stock ration, was a well balanced diet affording
rapid growth and high fertility*

Ration II was quite similar to the Steehbock

and Black No* 2965 ration containing a high amount of calcium in relation to
the phosphorus present, which in the absence of vitamin D produces severe rickets*
In order to determine whether the presence of vitamin D would prevent bent-nose,
to each 100 grams of ration II one gram of cod liver oil was added*
constituted ration III*

Realizing that 3

This

cent of Ca CO— might make the

ration so severe that the animals could not survive to 120 days, ration IV
was formulated with 1 per cent of Ca C0^ replaced by 1 per cent of yeast*
The presence of the yeast would cause more growth chiefly by stimulating the
appetite of the rats*

Ration V was identical with IV except that the yeast

was treated with ultra-violet light, thus adding vitamin D to the diet*

The

last two rations, VI and VII, were designed to be high in phosphorus in re­
lation to the amount of calcium present, the reverse of the other deficient
diets*

Since ration VI was quite complex and produced no striking results,

it was soon discarded and ration VII substituted*
Procedure
The technique employed was to keep all breeding animals on ration
I, the balanced stock ration* to prevent any difficulty in getting them to
breed*

Pregnant females were isolated in order to get accurate data on the

birth of the young*

The offspring were weaned at 28 days of age and put on

one of the experimental rations*
out the experiment*
Lansing is not high*

Shavings were used as cage bedding through­

Tap water was used, since its mineral content in Bast
Rations were kept before the animals at all times*

They were fed the experimental ration until at least 120 days of age, after

-12-

whlch they were etherized and their skulls examined to determine whether
or not they were bent*

Same of the bent animals showed the trait markedly

before they were 120 days of age, and these were transferred to the stock
ration and later used for breeding*
Three strains of rats were used, the bent stock and two control
stocks*

The animals constituting the bent stock were progeny of bent x bent

matings*

One control strain was the Wistar experimental strain derived from

animals obtained from the Wistar Institute of Anatomy*

The second control

strain consisted of black-hooded animals obtained from Dr* C* A* Hoppert*s
chemical laboratory at Michigan State College#

It was intended at the outset

that at least 50 animals of each stock would be reared on each of the seven
rations*

Due to a lack of animals, the Hoppert control stock was later

limited to ration II, the high calcium ration, and ration VII, the high
phosphorus ration#
Each animal used In experiment was given a number by a system
of ear markings*

He cords of matings were kept in order that the ancestry of

each animal could be traced*

Before any animal was discarded it was

etherized and the mid suture of its skull compared to a straightedge for the
final check as to whether the rostrum was straight or bent*

Ho final state­

ment on the state of the nose was entered In the record book before this
final examination was made*
Presentation and Discussion of Data
Growth Study*
One of the most effective means of determining the general response
of animals to a given diet is by studying their growth curves*

A number of

animals from the bent stock* subsisting on the various rations, were weighed

-13-

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200
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160
IV
120

£

100

•H

SO

VI
VII

III

II

20

28
Age In days
Bation I - stock ration
Bation II - high calcium ration
Bation III - high calcium ration plus cod liver oil
Bation IV - yeast, high calcium ration
Bation V - irradiated yeast, high calcium ration
Bation VI - high phosphorus ration
Bation VII - high phosphorus ration
GROWTH CURVES
Figure 3

-15-

at weaning time and later at 7 day intervals.

Their growth curves were

plotted (Figure 3)> and. the average weights are given in Table 1.

The number

used from each dietary group ranged from fourteen to twenty-eight, obviously
a fair sample of the whole 50 or more animals that were reared on each diet.
It is evident from these curves that while ration I, the stock
ration, supported the most rapid growth, ration II, the high calcium ration,
was the most severe on the animals.

On ration II their average weight actually

declined as much as 10.3 grams the seventh week, with a total gain of only
16.0 grams, as compared with an average of 118.0 grams gain for the animals
on ration I.

The average total gains of the animals on the other rations

ranged between these two figures, with no other curve showing a loss in weight*
The animals on ration III were consistently lighter than those on ration II
for the first four weeks.

It appears that this was due, in part at least, to

the repelling odor of the cod liver oil, causing the animals not to eat so
heartily.

The death of several of the animals on ration II after the seventh

week made it impossible to continue this curve, and consequently some of the
others also were discontinued after the seventh week.
Mortality Study
Few deaths occurred except on ration II, the high calcium ration.
Table II shows the mortality record of animals raised on this ration.
proportions dying were as follows?

The

1J.BG per cent of 112 rats of the bent

stock, 89.09 per cent of the 55 Wistar animals, and 7S.13 VeT cent of the
6H Hoppert rats*

-16-

Table II
MORTALITY OB' ANIMALS FED ON RATION II - HIGH CALCIUM RATION
Stock

No* of Animals
subject to
ration II

No* dead
before
120 day age limit

112

20

17*86 ± 2.HH

Wi'star stock

55

U9

89*09 ± 2*83

Hoppert stock

Sh

50

78*13 ± 3*1+9

Bent stock

Per cent
dead

These results tend to substantiate the work of Gowen^ in which he
has shown that the mortality rates differ among different strains of rats fed
on a ration deficient in vitamin D and high in calcium content*

These three

strains of animals differed in their reactions to the high calcium ration as
demonstrated by the mortality rates*

The mortality in the Wistar stock

exceeded that in the bent stock by 71*23±3*7^ Per cent; this difference was
19*03 times the magnitude of its probable error*

The mortality of the

Hoppert stock was 60*27=^*26 per cent higher than in the bent stock, the
difference being lU*15 times the size of the probable error*

Both these

differences were decidedly significant in a statistical sense*

The Wistar

and Hoppert stocks did not differ materially from one another, the former
exceeding the latter by only 10*96 J^*^9 per cent; this difference was only
times its probable error*
Seven, or 35 Per cent, of the animals of the bent stock which died
before reaching 120 days of age, were bent-nosed at death while only four,
or 0*72 per cent of the Wistar stock, and one animal, or two per cent of the
Hoppert stock showed the anomaly*

These figures, along with data presented

-17-

later in the paper, show that the strain which could survive the deficient
ration best produced the highest percentage of bent—nosed animals*
Deaths on the various other rations were too few to be of great
significance*

One rat among the animals of the bent stock on ration III

(high calcium plus cod liver oil) died before reaching 120 days of age*
animal displayed a normal nose*

This

The only other death in the bent stock was

on ration VI, the high phosphorus ration*

This animal also had a normal nose*

One bent-nosed and one normal Wistar animal died on ration IV, the yeast hi^i
calcium ration, and seven normal animals of the Hoppert stock died on ration
VII (hi^i phosphorus ration) before reaching the minimum age limit*

No

casualities occurred in any of the groups not discussed*
These results justified us in not using ration II (the high calcium
diet) in the genetic experiments to follow, because of its deleterious effect
upon the animals*

The errors introduced by the death of such high percentages

of animals would hamper genetic analysis*

From this standpoint alone any of

the other rations could have been used since the death rate of animals fed on
them did not exceed that normally expected for animals on almost any deficient
diet*
Study of rations in relation to -production of bent*
The cardinal data gathered in the nutritional work are given in
Table III*

The animals used in computing the per cent bent-nosed on each

ration Included only those which lived to 120 days of age or more*
Because of so many deaths in both the Wistar and Hoppert stocks
on ration II the ultimate numbers were too small to give conclusive data;

-18-

Table III
INFLUENCE OF THE SEVEN RATIONS ON THE FREQUENCY OF BENT-NOSE

Ration
Ration I
Stock ration

Bent stock:

Wistar stock

Hoppert stock

No* of
No*
io
animals bent bent

No* of
No.
$
animals bent bent

115

1

0*87

51

Ration II - High
Calcium ration

92

46

50*00

6

4

66*66

Ration III - High
Calcium ration
cod liver oil

48

5

14.17

54

0

0

Ration IV
Yeast, high
calcium ration

55

25

58

22

37-93

3*92

53

0

0

47*06

12

8

66*67

64.91

68

21

30.88

nation T
Irradiated yeast
high calcium ration

2

Ration VI - High
phosphorus ration

3^

Ration VII - High
phosphorus ration

58

37

No. of
No.
%
animals bent bent

l4

51

2

3*77

-19-

consequently these groups were not included in the discussion#

Likewise

the twelve Wistar animals on ration VI were omitted because of the paucity
of numbers*
A general survey of the different percentages of animals developing
bent-nose on the various diets shows that food must be a factor in determining
whether or not an animal shall develop the anomaly*

In the bent stock the

percentages vary from practically no bent-nosed animals in those fed on ration I
to as high as 6^*91$ bent-nosed on ration VII*

It was noted that one bent-nosed

rat on ration 1 was an under-sized individual*

The progeny of bent x bent

matings,, comprising the bent stock;, probably had about the same genetic constitution
as far as the bent-nose character was concerned*

Thus, with all environmental

factors approximately equal except for the food, one could unquestionably
attribute the variation in percentages of bent-nosed animals to the differences
in the diets*
The results for the Wistar stock roughly paralleled those for the
bent stock and pointed toward diet as a factor, but in no case was the incidence
of bent-nose as high as in the corresponding group of the bent stock*

On the

three rations, I, III and V, producing the lowest percentages of bending in the
bent stock, no bent-nosed animals appeared in the Wistar stock*

37*93 P®r cent

of the Wistar strain on ration IV were bent-nosed as compared with ^5*^5 per cent
of the bent stock, while only 30*6S per cent of the Wistar animals on ration
VII were bent as compared with 6^*91 per cent for the bent stock*

A little

discrepency has entered here; in the bent stock ration VII produced a higher
frequency of bent than ration IV while the reverse of this was true with the
Wistar stock, but as will be shown later the irregularity was hardly
statistically significant.

-20-

A more thorough analysis of the data revealed that the calciumphosphorus ratio was the decisive factor in the diet.

Eation II, containing

3 per cent Ca 00^, raised the incidence of bent-nose from less than 1 per cent,
as in ration I, to 50 per cent.

The addition of vitamin D in cod liver oil

(which tends to correct for the Imbalance in the calclunHphosphorus ratio) cut
the frequency of bent to 1*+.17 per cent, a decrease of 35.8 3 *. h.gg per cent
as compared with the frequency on ration II*
its probable error*

This difference was 7*35 times

The addition of yeast to the high calcium diet had little

effect on the Incidence of bent*

When the percentage of bent-nose among the

animals on ration IV was conpared to that among the animals on ration II, a
difference of *+*55 ^ 5*78 P©r cent was found, which was only 0.S0 times its
probable error.

A difference this great might be expected as ration IV had only

2 per cent Ca CO^ while ration II contained 3 P®r cent Ca 00^.

A very marked

difference in per cent of bent was again observed when vitamin D was added in
ration V.

This time it was in the form of Irradiated yeast.

Irradiating the

yeast alone reduced the frequency of bent from *+5*^5 Per cent on ration IV to
3*92 per cent on ration V, a difference of *+1*53 * *+.SS per cent which was
S.51 times its probable error*
The percentage of bent obtained on ration VII indicated that reversing
the calcium-phosphorus ration, making phosphorus high, tended to increase the
frequency of bent-nose*

This was not borne out with the results from ration VI*

Although rations VI and VII were both high in phosphorus, VI was not as
imbalanced as VII, since it was a more varied diet and contained 5 Per cent
milk powder which would tend to raise the calcium content*

21-

Statistical analysis did not reveal a striking difference between
the results from the high calcium rations and those from the high phosphorus
rat ions • Hation VII gave a percentage 1^*91
of ration II*

5*^9 per cent higher than that

This difference was only 2*72 times its probable error*

Hation VII gave 19* H6

6*IS per cent more bents than ration IV, and this

difference was 3*^5 times its probable error*
differed by 17#85

The two high phosphorus rations

7*15 Per cent* a difference 2*50 times as great as its

probable error*
Thus it could be concluded that the calcium-phosphorus ratio was a
factor in predisposing bent-nose*

On a ration in which these elements were

quite well balanced practically no bent animals developed*

When young rats

were fed a ration in which this balance had been disturbed, the frequency of
bending increased*

The introduction of vitamin D corrected for the imbalance,

and the frequency of bending diminished*

These conclusions were likewise borne

out by the results from the Wistar animals*
Comparison of growth with incidence of bent-nose*
When a comparison was made between growth rate and the incidence of
bending on the various diets among the animals of the bent stock, it was noted
that in general those rations producing slight growth caused a high frequency
of bending*

The contrary was likewise true*

Ration I which produced the

greatest gain (118 grams) in seven weeks produced less than one per cent
bent-nose*

In contrast of these figures, animals gained only 16 grams in

seven weeks on ration II, while the frequency of bending on this ration was
50 per cent.

The ration producing the highest frequency of bent-nose, ration

VII, cause the animals to gain on an average only ^9*1 grams*

Ration IV which

produced a rather high frequency of bent-nose (^***5 Per cent) produced an

-22-

Table IV
THE EFFECT OF EACH RATION ON THE FREQUENCY OF BENT-NOSE AS COMPARED
WITH ITS EFFECT ON GROWTH RATE
Ration

per cent
Gain in weight daring
bent-nosed
tbs weeks after
,
_____________ ________ ________________ weaning_________ _

I - stock ration

0*87 per cent

118*0 grams

V - irradicated yeast,
high calcium ration

3*92 per cent

96*7 grams

III - high calcium
ration plus cod liver oil

lH*17 per cent

U7.5 grams

IV - yeast, high
calcium ration

^5*1+5 per cent

76*0 grams

VI - high phosphorus ration

U7#o6 per cent

59*3 grams

II - high calcium ration

50*0 per cent

16.0 grams

VII - high, phosphorus ration

6U.91 per cent

U9.I grams

-23-

average Increase in weight of JG grams for the seven weeks*

While vitamin D

when added to ration IV, forming ration V, decreased the percentage of bentnose to 3*92 per cent, It increased the average gain in weight to 96*7 Sran®*
Ration III was quite irregular.

The frequency of bent-nose among those

animals oh this ration was 1^*17 per cent, but the animals gained on an average
only ^7*5 grams in the seven weeks*

However as before stated it was difficult

to get the animals to eat much of a ration containing cod liver oil, and this
was probably responsible for this irregularity.
Irregular.

Likewise ration II was

However, the total gain in weight of animals fed ration II varied

from that in the other groups in that the animals actually lost weight during
the last two weeks*
Bone-ash studies
In this investigation it was desirable to make bone ash determinations
for two reasons;

(l) to determine the severity of the rickets induced by the

deficient diets, and (2) to know whether the different strains behaved differently
toward the same ration*

The percentages of ash in the bones are given in

tables V to IX inclusive*
To makes these determinations groups of animals from the three
stocks were sacrificed after they had been fed on ration II (high calcium
ration) for twenty, forty, and sixty days, and on ration VII (high phosphorus
ration) for twenty and forty days*

The animals were etherized and their

femurs were removed, after which all the surplus tissue was removed by rubbing
with cheesecloth.

Fat was extracted with alcohol in the conventional manner,

and the bones were dried and weighed*

After this they were ignited in an

electric furnace to destroy the organic matter; the ash was then weighed, and
the percentage of bone ash was calculated*

-2^-

The per cent of inorganic matter in bone has for a long time
been the principal criterion for determining the severity of rickets in rats*
Femurs of normal rats will vary from the extreme low of ^5

as high as 60

per cent ash, depending somewhat on age, sex, food and various other factors*
Females tend to have a higher ash content than do males, and as animals grow
older the ash content of their bones normally increases slightly*

A proper

calcium - phosphorus ratio in the diet, with ample vitamin D, tends to produce
a maximum percentage of ash*

However, during pregnancy or lactation the percentage

of bone ash tends to decrease due to the drain on the inorganic elements of
the mother for the formation of the embryo or the production of milk*

This

percentage of bone ash is markedly lower than normal in animals suffering
from rickets*

Often it will be as low as 30 per cent, or even lower,

depending

■c^on the severity of the condition.
While both rations II and VII produced rickets, the high calcium
ration brought about a much more extreme type than the high phosphorus ration*
This was probably due to the fact that the ca!cium-phosphorus ratio is much
more Imbalanced in the high calcium ration than in the other*

The high amount

of calcium in ration II was obtained by adding 3 Per °ent of Ca CO^ while the
high phosphorus ration was developed merely with a high grain content and
no milk powder*

For the most part the percentages of bone ash on the high

calcium ration ran below 30 per cent with an extreme low of 19*5^ Per cent*
These percentages were probably bordering on the minimum
with survival.

limit compatible

The difference in time beyond the 20 day period which these

animals were fed this ration produced no significant difference in percentage
of bone ash*

-25-

The high phosphorus ration caused the percentage of hone ash
in most cases to drop within the rachitic range, i.e. around ^0 per cent or
lower, although in no case was the percentage extremely low*

The Wistar

animals which had been on this high phosphorus ration for forty days were
on the border line between rachitic and normal animals*

All three stocks

showed an increase in percentage of bone ash of the ^0 day period over the
20 day period*

This indicates that the physiological mechanism of these

animals had begun to adjust itself to the imbalanced calcium-phosphorus ratio*
In general it can be stated that no stock was more resistant or
susceptible to the deficient diets than were the other two stocks in regard
to percentages of bone ash*

In various groups the average bone ash percentage

of one strain may have appeared higher or lower than those of the other two,
but with no stock was the percentage consistently hi^i or low throughout.
Seven of the animals fed forty days on ration II were bent-nosed
when etherized*

It is interesting to note that these animals were neither

lower nor higher in bone ash content than other members of their group*

This

would show that the factors for bent-nose are specific rather than influencing
the reaction of the whole body to the deficient diet*
It is interesting to compare the results of the bone-ash
determinations with those of the mortality study in regard to the resistance
to the deficient diets displayed by each of the three strains*

While the

death rate showed that the bent stock was superior to either of the other
two stocks in resisting the deleterious effects of ration II, the strains
did not differ materially in percentage of bone ash.

-26Table V
■PER CENT OF BONE ASH IN THE FEMURS OF ANIMALS
FEB 20 BAYS ON RATION II - HIGH CALCIUM RATION
Bent Stock

Wistar Stock

Animal No*

$> Ash

3U06

2U.67

S 3327

2 S.90

tf 3l<07

30*97

9 3328

<? 3U08

26.15

<7 3 ^ 9

tt

Animal No.

# Ash

Hoppert Stock
Animal No*

$ Ash

26.07

e 3057
<s 3058

29.99

5 3329

27.80

<? 3059

28.39

27.0^

9 3330

26.80

^ 3060

26.7U

3U10

29. SI

9 3331

29.U9

9 3062

26.77

ef 3^11

29-37

9 3332

31.05

9 3063

28.96

<f 3^12

27-75

9 3333

32.63

a

306U

31.79

9 3^13

2S.03

9 333^

30.72

<t 3065

3U.U3

9 3U1U

30*09

9 33*+3

33.U0

5 3066

32.69

5 3^15

29.02

9 33^

32. Us

9 3067

29.50

9 33^

3U.19

9 3068

29.96

9 33U6

31-57

9 3069

33.70

9 33^7

29.67

9 3070

31.55

9 33^8

33-79

9 3071

28.60

Ave*

30.68

Ave*

30.39

Ave.

2S.29

32. Ui

-27-

Table VI
PER CENT OF BONE ASH IN THE FEMURS OF ANIMALS
FED UO BAYS ON RATION II - HIGH CALCIUM RATION
Bent Stock
Animal No*

Hoppert Stock

Wistar Stock
$ Ash

Animal No*

$ Ash

Animal No*

Ash

20.02

cT 30UU

29.^3

25*07

e 3311
* e 3312

21*58

e

30U5

27.38

269s

25 *S7

<7 3313

22.US

9 30U7

2U.31

9 2699 .

25.31

•

e

33iU

21*50

9 30US

23.90

9 3U01

2U.97

* <3 3316

23.19

9 30U9

27.82

9 3U02

23.76

9 3317

21*19

<3 3051

25.89

9 3^3

2U.59

• 9 3318

21.23

<7 3052

27.73

9

2U.29

9 3320

21.09

<3 3053

31.72

9 3^5

25.05

<3 3321

21*50

cT 305U

28.91

• <3 3322

20 *9S

9 3055

2U.U8

* <3 3323

20*93

9 3056

23.36

<3 332U

20*73

Ave*

21.37

Ave*

26.81

<*• 269U

25 .9 S

* <#• 2695

e

Ave*

2U.99

♦Bent—nosed animals

-28-

Table VII
PEE CENT OOP BONE ASH IN THE FEMURS OF ANIMALS FED
60 DATS OF RATION II - HIGH CALCIUM RATION
Bent Stock:
Animal No.

$ Ash

Hoppert Stock

Wistar Stock
Animal No.

Ash

Animal No.

$ Ash

2678

25.13

9 3335

26.57

e

3039

30.95

2679

19.53

9 3337

23.65

a

30^1

39.6^

«f 2600

22.66

9 3338

25.93

<* 3038

26.26

a

26si

24.30

9 331<0

22.U1

e

30*10

38.05

5 2682

25.46

9 33^1

20.72

9 30U2

27.27

9

26SH

27.00

9 30^3

33.56

9

2686

21.31

9 2687

21.45

Ave*

23.36

e

Ave.

23.86

Ave.

32.62

Tabic* VIII
PER CERT OF BORE ASH IN THE FEMURS OF ANIMALS FED
20 DAYS ON RATION VII - HIGH PHOSPHORUS RATION
Bent Stock

Wistar Stock

Hoppert Stock

Inimal No*

$ Ask

Animal No.

<? 3^39

35.16

<f 35-72

Us.85

e 3099

39.^8

<r 3UU0

31.81

rf 35-73

36.71

e U301

35.63

3UU1

31.60

rf 35.7^

U1.17

5 U302

32.02

ti> 3UU2

33.33

«f 35.75

UU.17

? U303

38.05

S 3^3

32.9S

<f 35.76

U1.06

a 3093

3U.U5

5 3UUU

37.00

rf 35.77

U2.09

€ 309U

36.15

e 35-78

U1.77

ef 3095

36.U2

<r 35.79

38.96

d 3096

37.20

<f 35-80

U1.00

«f 3097

35. ^

35.81

36.30

<f 3098

37.06

5 35-82

Ul.Us

Ave*

U0.96

Ave*

36.U0

Ave*

33*66

$ Ash

Animal No*

# Ash

-30Sable IX
FEB CENT OF B O M ASH IN SEE! FMJRS 0? ANIMALS
FED ^

DAYS ON RATION VII - HIGH PHOSPHORUS RATION

Bent Stock:

Wistar Stock

Animal No*

$ Ash.

if

3*32

39-20

if

<f

3*33

if

Hoppert Stock
Animal No*

$ Ash

Animal No*

# Ash

3566

H5.11

ef JOSH

H3.16

H0.09

9 356s

H7.21

«? 30S5

H6.31

3H3H

39.72

9 3569

H9.09

d- 3086

H5 . 2H

9 3^35

Ho.3o

9 3570

H7.36

<T 3087

H2.37

9 3^36

H2.6o

9 3571

H6.H2

<? 3088

H3.60

9 3>+37

H2.61

9 3090

37-30

9 3^38

H3.07

9 3091

36.H7

9 3092

38.99

Ave*

Hi. 68

Ave*

Hi .08

Ave*

H7.0H

-31-

Results of subjecting animals to a deficient ration at 16 days of age*

An attempt to increase further the incidence of bending among the
progeny of bent x bent matings by putting the animals on the deficient ration
at an earlier age proved to be futile*

Forty rats were used in this test*

At

the age of sixteen days the animals with their mothers were put on ration II
(high calcium ration) •

They were weaned at twenty-one days of age, the mother

being returned to ration I (stock ration), but the young were continued on the
deficient diet*
Such treatment was too severe for the young animals, and they all
died long before they reached an age of 120 days.

As seen in table X, seven

of the animals died at 28 days, and the remainder died at ages ranging from
fifty to seventy-eight days.

The seven that died early were all of one litter,

indicating that they were more susceptible in some way than the others*
It was very evident that this plan of feeding could
in a

not be followed

breeding experiment as most of the animals would be dead before they had

reached the age at which the bending normally appeared.

However, three of

these animals did develop bent-nose, as was discovered on examination of
their skulls when they died.

One of them was 60 days of age, one sixty-one,

and the third seventy-six days old when it died*

Results of inbreeding attempt*
An inbreeding experiment was started to determine whether inbreeding
with selection would increase the frequency of bending*

If bent-nose was

caused by multiple factors or by a simple recessive or simple dominant gene
with modifying factors a significant difference should have been observed
between the first and second generations of inbreeding, in which only animals
with a high degree bending had been used in the matings*

-32-

Table X
AGES AT WHICH PROGENY OF BENT X BENT MATINGS DIED WHEN PUT OH
RATION II (HIGH CALCIUM RATION) AT SIXTEEN BAYS 07 AGE AND
WEANED AT TWENTY-ONE BAYS 07 AGE.
No. dying at the given age.
28 days of age

7

50 days of age

2

5^ days of age

3

55 days of age

1

56 days of age

2

57 days of age

1

5S days of age

1

59 days of age

1

60 days of age

2

6 l days of age

5

62 days of age

l

6 U days of age

1

67 days of age

l

69 days of age

2

70 days of age

1

71 days of age

3

73 days of age

1

75 days of age

1

76 days of age

1

77 days of age

2

7S days of age

1

Total no* of animals

UO

-33-

Fortunately four fertile bent-nosed females and one fertile bent­
nosed male, all of the same litter were obtained*

These were progeny of a

bent x bent mating and had been reared on ration VII (high phosphorus ration).
The four bent—nosed females were mated to their sib producing nine litters
consituting a total of seventy-six offspring*
At that stage of the experiment, data indicated that ration II high calcium ration produced the highest frequency of bent-nose.

For this

reason these inbred animals were subjected to ration II at weaning age —
twenty—eight days*

But the inbred animals did not survive the bad effects

of this ration as well as animal s of the bent-stock had*

One hundred and

twenty days after birth, or after 92 days on the deficient feed, only 25, or
32*9 VeT cent of the animals were yet surviving*

Of the bent stock 82.lU

per cent survived the effects of this ration to the 120 day age limit*

This

decrease in vitality may have been due in part at least to the inbreeding*
Among the twenty-five animals living 120 days, seven, or 2S per
cent were bent*

This is a much lower percentage than 50 per cent, that

obtained in the bent stock on the same ration mentioned in the previous
section.

However, twenty-five animals were too few to furnish significant

results*

Of the fifty-one animals dying before they were 120 days old,

sixteen, or 31*37 per cent were bent.

These figures would indicate that

neither bent nor normal animals tended to die more readily within the 120
day period.

The data are given in table XI.

It was impossible to rear a second generation in this phase of
the experiment*

This was due to the high mortality and low fertility of the

animals that lived to 120 days of age.

So few fertile animals matured that

brother sister matings could not be made*

- 3^
Table XI
FREQUENCY OF BENT-NOSED ANIMALS AMONG- THOSE OF THE
FIRST GENERATION OF INBREEDING.
No* of
Animals
Animals living 120
Animals not living 120 days

25
51

No.
Normal

No,
Bent

Per cent
Bent

IS

7

2S.00

35

16

31*37

However* these animals were not all reared in vain*

Each animal

was weighed at birth, marked* and again weighed at weaning time*

It was

desirable to know whether there was any correlation between size at birth
or gain in weight during early life* and tendency toward bent-nose*

Had

it been found that animals which later developed bent-nose were smaller at
birth than normals, one would have suspected some prenatal factor as having
a causative influence on bent-nose*

Moreover* the fact that bent—nosed

animals did not gain as rapidly before weaning might have led to the
discovery of some factor operating early in life*
Data presented in tables XII andXIII show that there was no
significant difference between the mean weights at birth of bent and normal
animals*

The same is true when the mean weights at weaning age and the

gain in weight from birth to weaning are compared*

The difference between

the means in birth weights was *^3 ±. .88 grams, or *U$ times its probably
error*

The means of the weight at weaning time differed by U.25 ± 9*0^

grams* a difference of *1*7 times its probably error*

The difference between

-35-

the means of gain in the two groups of animals was only .42 times its
probable error, it being 3*68 ± 8*63 grams,

Thus, there is no evidence

that the factor or factors causing bent-nose affect body weight either before
or after birth*
It has been shown that food affected the incidence of bent-nose
and also the growth rate, and that low growth rate tended to be correlated
with a high incidence of bending.

However, the data Just presented indicate

that the high frequency of bent-nose was not a result of low growth rate
nor was the reverse true.

In the animals of the inbred experiment, having

approximately the same genetic constitution for bent-nose, practically no
difference was observed between the weights and growth rates of the bent
animals and those of the noxmals.

This would tend to Indicate that the

dietary factors causing a high frequency of bent-nose ware different from
those affecting a low growth rate*
Table XII
COMPARISON OP MEAN WEIGHTS OP BENT-HOSED ANIMALS
AND NORMAL ANIMALS OP THE INBREEDING EXPERIMENT
Normal animals
_________
Mean wt.
at birth

Mean Wt*_______ <T

P.E.

Mean Wt.

<T

P*E*

.92

.62

5*97 ©as*

,
*94

*°3

39*33 S®8*

9*92

6.69

35*08 gns.

9*02

6.08

38.79 ©>“>•

9*71

6*55

29.11 ©»»•

8.77

5*92

6*4o gms.

Mean wt.
at weaning

Bent animals

Mean gain
in weight

-36Table XIII
WEIGHTS 07 N0BM4L ANIMALS IN
INBBEEDING EXPERIMENT
Animal Ho.

Wt. at birth
in grams

Wt. at weaning
in grams

Gain
in grams

rf 2802

6.70

33.**o

26.70

9 2805

7.20

3^52

27.32

5 2807

7.1*4

35.30

28.16

5 2827

6.50

17.30

10.80

cT 2828

6.80

11.30

*4.50

9 2830

7.00

2*4.60

17.60

e 2631

7.*40

25.80

18.*40

5 2832

6.90

21.00

1*4.10

? 2833

7.*40

27.90

20.50

<i 2810

5.25

32.70

26.U5

281*4

5.2*4-

32.00

26.76

9 2816

5.1*4

32.50

26.36

<f 2835

7.80

36.20

2S.*40

9 2836

6.80

3*4.*40

27.60

e 2837

7.70

5*4.30

*46.60

9 2838

6.00

31.30

25.30

? 2839

7.50

*42.60

3*4.10

9 28*40

7.00

35.70

28.70

9 28*41

7.70

*46.30

38.60

2861

6.80

38.10

31.30

9 2862

7.60

5*4.00

*46.*40

-37Table XIII (continaed)
WEIGHTS OF NORMAL ANIMALS IN
INBBEEDING EXPERIMENT
Animal No*

Wt. at birth
in grams

Wt* at weaning
in grains

Gain
in grams

$ 2864

6.80

37.05

30.25

rf 2865

7.80

52.70

44.90

g 2866

7.70

51.80

44.10

9 2867

7.00

ItS.80

4i.S0

9 2868

7.20

U9.8O

42.60

tf 2869

8.1(0

55.60

47.20

e 2818

5.68

36.00

30.32

9 2819

5.20

33.00

27.80

9 2821

5.76

47.00

41.24

2822

6.35

44.00

37.65

a 2823

5.87

39*50

33.63

9 2824

5-72

U7.90

42.18

<? 2825

5.05

33.50

28.45

2826

5*75

1(5.50

39.75

2852

5.60

41.10

35.50

9 2853

5.50

41.80

36.30

2S54

5.10

27. 80

22.70

rf 2857

5.70

27.60

21.90

9 285S

5.50

39.00

33.50

2844

5*70

46.80

41.10

9 2846

5.20

40.20

35.00

-38Table XIII (continued)
WEIGHTS OP NORMAL ANIMALS IN
INBREEDING EXPERIMENT
Animal No*

e

Wt* at birth
in grams

Wt* at weaning
in grams

Gain
in grams

2SU7

5*70

4U.1#)

35-70

9 2SUg

5.22

Ul.70

3 5 .^

e

28^9

5.62

U2.70

37.O8

9 2850

5.05

37.20

32.15

e
a

2S51

5.60

33.20

27.60

2870

7.60

5^.70

^7.10

9 2871

6.20

U7.6O

Ul.lio

<? 2872

7.30

5U.OO

U6.70

9 2873

6.00

U5.8O

39.80

9 2871*

6.U5

U3.OO

36.55

9 2876

6.50

52.140

H5.90

-39Table XIII (continued)
WEIGHTS Off BENT-NOSED ANIMALS
Off INBREEDING EXPERIMENT
Animal No*

Wt* at birth
in grams

Wt* at weaning
in grams

Gain
in grains

a 2801

6 .7 5

3 2 .7 3

2 5 .9 8

t! 2803

6 .2 6

3 0 .3 0

2 4 . o4

9 2804

6 .8 0

32*85

2 6 .0 5

<7 2806

6 .3 5

3 2 .7 7

2 6 .4 2

2808

7 .5 3

3 4 .0 0

2 6 .4 7

2829

7 .6 0

2 0 .0 0

1 2 .4 0

rf 2834

6 .9 0

2 8 .7 0

2 1 .8 0

9 2809

4 .7 3

3 2 .3 0

2 7 .5 7

<7 2811

5 .2 3

33*20

2 7 .9 7

<7 2812

5 .5 5

3 4 .3 0

2 8 .7 5

<7 2813

5*27

3 1 .0 0

2 5 .7 3

9 2815

5 .0 0

30. 4o

2 5 .4 0

«7 2817

4 .6 0

2 4 .6 0

2 0 .0 0

9 2842

6 .6 0

4 7 .5 0

4 0 .9 0

9 2843

7 .0 0

4 7 .2 0

4 0 .2 0

<7 2863

7 .0 0

5 4 .4 o

4 7 . 4o

<7 2820

5 .4 4

4 3 .0 0

37.56

9 2855

4 .9 0

2 3 .1 0

1 8 .2 0

9 2856

5 .2 0

3 4 .4 o

2 9 .2 0

9 2859

4 .6 0

2 6 .9 0

2 2 .3 0

9 2860

5 .7 0

3 4 .4 o

2 8 .7 0

9 2845

5 .6 0

4 5 .0 0

3 9 .4 0

9 2875

6 .8 0

5 4 .0 0

4 7 .2 0

-ijo-

Summary of Nutritional Studies
1*
in the rat.

Diet is a factor influencing the manifestation of 'bent-nose
With other factors controlled* different diets produce different

frequencies of bent~nose*
2*

The calcium-phosphorus balance in the food is a very important

factor in determining whether bent—nose develops*

That is* if the calcium—

phosphorus ratio of the diet is imbalanced* bent-nose occurs*

This imbalance

may be affected by adding either too much calcium or too much phosphorus*
3*

As the introduction of vitamin D ameliorates the deleterious

effect of the disturbed calcium—phosphorus ratio* it likewise decreases the
frequency of bent-nose*
U*

In general* the percentage of bent-nose produced by the

various rations varied inversely with the growth rate*

That is* the rations

most deleterious to growth favored the development of bent-nose*

On the

same ration, however, significant differences in weight or gain in weight
were not found between the bents and their normal siblings*
5*

The bent-nosed strain was more resistant to the high-calcium

ration as judged by the percentage of mortality than was the Wistar strain
or the Hoppert strain*
6*

Bone ash determinations failed to demonstrate any difference

in bone ash content between the three different strains when fed on a
deficient ration*

Ration II (high calcium ration) produced a lower

percentage of bone ash than did ration VII (high phosphorus ration)*
7.

The effect of putting the animals on ration II at the age of

sixteen days and weaning them at the age of twenty-one days could not be
determined, because they were unable to withstand the treatment and died
before showing the bent-nose character*

-Ul-

8*

An attempt to increase the frequency of bent-nose by inbreeding

was unsuccessful, for the progeny reared on the deficient ration showed low
fertility and high mortality*
9*

We were unable to prove the existence of prenatal or postnatal

factors in the production of bent-nose which influenced the birth weights,
weights at weaning age, or increase in weight of the bent-nosed rats*
10*

Since ration VII (high phosphorus ration) produced the highest

percentage of bent-nosed animals and yet did not kill the animals before they
were 120 days old, it proved to be the most desirable ration on which to rear
the animals in the genetic experiments*

-1*2GENETIC FACTOBS CAUSING BENT-NOSE
Introduction
9
Work like Gowen*s

may prove very enlightening in the study of

inheritance of hone abnormalities, since often nutritional factors are
involved*

He demonstrated that separate lines of rats showed distinct

differences i*1 respect to survival on a diet deficient in vitamin D and high
in calcium.

The author has data tending to substantiate this.

"Probably the most satisfactory genetic explanation of a bone
i £ op

abnormality in rodents is that given by Hunt, Mlxter and Permar
worked on flexed tail in the mouse.

who

This was a condition in which the

caudal vertebrae were malformed and fused, usually producing varying degrees
of flexure in the tail.

Although it has been definitely shown that this

trait is inherited as a recessive, modifying factors are involved.

The

19
development of this character has been further studied by Kamenoff.
A condition similar to flexed tail in Mus muscuius has been
reported by Huestls and Bar to

in Peromyscus.

It was stated that this

character appeared to be due to two recessive genes.
12
Green and Brownv, in making a study of hereditary variations in
the skull of the rabbit,showed that profound variations in the form of
the skull could be transmitted unaltered from parent to offspring.

However,

no definite genetic scheme was advanced to explain the inheritance of this
variation.

A genetic cranometric study of two species of mice by the late

0. V. Green11

shown that the size of the skull tends to be inherited.

However, Green states that a thoroughly satisfactory genetic explanation

-43-

appears impossible with the data he has gathered#

Colton*^ reported woric

on the Hhigh-browM rat, but this study failed to demonstrate whether the
character was inherited#
Procedure for Studying the Genetic Pactors Causing Bent-no3e in the Rat#
Before the genetic experiments could be undertaken, a suitable
ration had to be found#

As mentioned in the preceding section, there were

two requisites for this ration*

(l) it could not be so severe as to kill

the animals before they reached 120 days of age, and (2) it should produce
the maximum incidence of bents among progeny of bent x bent matings without
causing too high a death rate#
Since it was observed early that ration II (high calcium ration)
produced a high incidence of bending it was first employed in the breeding
experiment, but when more data were gathered the high mortality under ration
II was revealed, and it was discarded for ration VII (high phosphorus ration)#
When all the data on the different rations were analyzed, it was found that
ration VII filled both of the above requisites#
A strain of rats genetically free from factors for bent-nose was
needed for crossing with the bents#

The Wistar experimental strain and the

Hoppert strain were tested for this purpose*

The data given in table III

show that the Wistar experimental strain carries factors for bent-nose to
a relatively hi^i degree*

On ration VII, 30.SS per cent of the Wistar

animals were bent as compared to 6*+#91 P©r cent bent among those of the
bent strain*

This reduced the probability of finding a strain genetically

free from bent-nose since so many strains of rats were related to the Wistar
stock#

Dr# W. E# Castle states it was quite probable that our red-eyed-yellow

-UU-

strain formerly Obtained from him, in which the anomaly first appeared and
from which our bent stock was developed, were related to the Wistar animals*
0?he strain was originally secured in the United States from a fancier in
England, but in the course of subsequent experiments has been crossed with a
number of strains, among which may have been animals from the Wistar Institute*
!Ehe Hoppert strain proved to be nearly free from bent factors*
only 3*77

cent of this stock was bent*

On ration VTI

Since the Wistar stock carried

factors for bent-nose, it was highly probable that the Hoppert stock was as
free from bent-nose as any available strain*

Dr. Hoppert*s animals were

originally obtained from the University of Wisconsin, beyond which their
ancestry was unknown.

One other alternative would have been to develop a

strain free from bent-nose by inbreeding and selection, but this would have
extended the duration of the experiment beyond possible bounds.

For these

reasons the Hoppert stock was chosen for the P^ matings*
All breeding animals were kept in ration I, the balanced stock
ration.

Pregnant females were isolated, and the date of the birth of the

young was recorded.

At twenty—eight days of age the young were weaned and

transferred to the deficient ration, the mother being returned to the breeding
cage.

The deficient ration was kept before the animals constantly.

After

they were 120 days old they were etherized and the mid sutures of their skulls
were carefully compared to a straightedge to determine whether they were
bent or normal*
P

matings were of bent animals of the bent stock mated to normal

animals of the Hoppert stock.

Since the deficient diets rendered the animals

of little use for -breeding, ample F-^s were reared on t'ne stock ration for

-H5-

maklng the

x

matings.

bent-noBed individuals.

These 3^ animals were also back-crossed to

Normal animals, the progeny of bent x bent, were

mated to 'bents in an attempt to determine whether or not they carried the
factors predisposing to bent-nose.

Also* brother-sister matings of bents

were mads in order to study the Change in incidence of bending afforded by
inbreeding, as reported in the previous section*
Presentation and Discussion of Data
Data obtained in nutritional studies
Before making an analysis of the percentages of bent-nose among
the F^, Pg, and back-cross generations it was well to analyze genetically
the data obtained in the nutritional studies,

These data are given in

table XIX*
The figures secured in that study furnish evidence that bent-nose
is influenced by genetic factors.

When a group of animals were fed on one

particular ration with other environmental factors constant, only a part of
these developed the bent-nose condition.

For instance, of the animals of the

bent stock fed on ration VII, only 6U.91 per cent were bent-nosed.

Thus,

there must have been some innate factors acting toward the manifestation of
bent-nose in these animals which were absent, or in some way deficient, in
the remaining 35*09 per cent.
to be bent-nosed or all normal.

If not, one would expect all of these animals
The same reasoning holds wherever both bent

and normal animals appeared in a group fed on any of the diets*
However, all the groups of the bent stock on the various rations
probably had approximately the same genotypes for bent-nose.
different rations induced different percentages of bents.
genetic constitutions react differently to different diets.

Yet the

Thus, certain
An animal

-U6possesslng the genes which would cause it to appear bent on the high
phosphorus ration might be normal if fed one of the other diets.
The fact that the three different stocks, when fed on the same
ration with other environmental factors nearly constant, produced different
percentages of bent—nosed animals is evidence that the character is genetic#
The percentage of bent-nosed animals of the Wistar stock on ration VII was
30*88 per cent as compared with 6^.91 per cent for the bent stock on the
seme diet*

The difference between these percentages was 3^*03 ^ 5*^6 per

cent, a difference which was 6*01 times its probable error.
stock on the same ration 3*77 P®** cent were bent-nosed.

In the Hoppert

This was 6l.ll*

U.58

per cent less than that of the bent stock, a difference 13*35 times its probable
error, and 27.11^^*18 per cent less than that of the Wistar stock.
latter difference was 6.H9 times its probable error.

The

Thus, it was evident

that there was a difference in the innate constitution of these different
stocks as to the production of bent-nose.

Factors for bent-nose must have

been rare in the Hoppert stock, more prevalent in the Wistar stock, and
still more numerous in the bent stock.
Only normal-nosed animals were used for matings in the Wistar
stock, and for this reason one would expect a greater variation in the
accumulation of factors for bent-nose in the Wistar stock than in the bent
stock in which all matings were bent x bent.

Since they were all normal,

it was not known which of the normal Wistar animals used for breeding
carried genetic factors for bent.

This probably explains why, while ration

IV produced a lower percentage of bent-nose than did ration VII in the
bent stock, a higher percentage of bent-nose appeared on ration IV

-^7-

than on ration VII in the Wistar stock#

The parents of those Wistar animals

on ration IV may have carried more genes for bent-nose than did the parents
of those on the other ration#
Data obtained in genetic studies
The data secured from the various crosses

hack crosses*

etc#) do not reveal any simple mode of inheritance for bent-nose#

Although

several theories might have been advanced as to the factorial situation for
bent-nose, none were found that satisfactorily fits all the ratios obtained.
However, the data are quite enlistening#

See table XIV#

One of the characteristics of the data given in table XIV which
first appears to the observer is the decline in percentage of bent-nose with
a decrease of bent-nosed ancestry#

Progeny of bent x bent matings were

6H»91 p©r cent bent-nosed, while progeny of bent x normal (from bent x bent)
matings were only 29*07 per cent bent-nosed#

Farther down the scale bent

x normal (Hoppert stock) matings gave progeny of which only 3*36 per cent
were bent-nosed#

With all environmental factors constant, such data

afford evidence that the character is genetic#
Unquestionably there is evidence of segregation#

There were only

3*36 per cent bent-nosed among the F^ animals from bent x normals (of the
Hoppert stock), and of the

from the matings F^ x F^, the F^’s derived

from matings bent x Hoppert normal, 10#

per cent were bent#

Although

this was not a high percentage of bent-nosed animals in the F^ generation,
it was significantly higher than that obtained in the F

generation#

The

difference was 7#35 ± 1*96 per cent, which was 3#75 times its probable error#

-te-

Llkewise, there was a significant difference 'between tixe percentage of
bents among the F^*s and that among the backcross (J1 x bent) progeny.
This difference was 7*^9
probable error.

2*63 per cent which was exactly three tizzies its

The percentage of bent among the backcross progeny was low,

and constituted the greatest barrier in formulating any genetic interpretation.
The data indicate that bent-nose is recessive to normal.
percentage of bent among the

The

animals was very much lower than that expected

if dominant factors were responsible for the trait#

If the factors were

dominant one would expect between 50 and 100 per cent of the P^*s to be
genetically bent and there are no good reasons for supposing that the action
of ration VII would have lowered the phenotypic percentage to 3*3^ per cent.
The more logical explanation is that the factors are recessive and that the
Hoppert stock carried some of these factors for bent-nose#

Bent-nose genes

were present in the Hoppert stock as Indicated by the two bent nosed Hoppert
animals which appeared in the nutritional experiments#
It appears that more than one pair of genetic factors are responsible
for bent-nose.
mouse?^,^ ,^

However, as with Reed*s investigation on harelip in the house
it may be that one gene is necessary for the production of the

character and the others are merely modifiers.

If the character were

controlled by two recessive genes one would expect 6.25 per cent of bents
in the

generation.

This is not beyond the bounds of probability when

compared to the 10.71 p®r cent actually obtained, the difference being
3#U6 ^ 1.6l per cent, which is only 2.15 times its probable error.

However,

such a genetic scheme would not give a backcross percentage comparable to
that obtained.

One would expect 25 J?eT cent bent-nosed in the progeny of

the backcross, and actually 11.25 per cent was all that appeared.

Three

-1Q-

Table XIV
Sm^JENCIES OF BENT-NOSE AMONG ANIMALS FROM THE
VARIOUS CROSSES, ALL REARED ON RATION VII
Offspring
Hrpe of mating

Total
offspring

Number
bent-nosed

Bent x bent

58

37

6H.91 ± *±.22

Normal (from bent x bent)
x bent

86

25

29.07 ± 3 .3 7

119

b

168

18

10.71 ± i*6l

80

9

11.25 ± 2.3s

F^ (bent x normal)
\

tsirf x Fis)

Backcross (F^ x bent)

Per cent
bent-nosed

3.36

*■

1-11

-50-

pairs of recessive factors would give 12*5 P©r cent bent-nosed among the
backcross progeny, which closely approximates the percentage obtained*
However, among the I*2 animals,a percentage of 1.6 would be expected.

Thus,

the actual percentage of bent-nosed animals in the Fg generation is too high,
to Justify this theory.
Another type of mating, the results of which are well worth further
consideration, is that of normal animals (from bent x bent matings) mated to
bent animals.

Such matings were desirable in order to test

such normal animals carried factors for bent-nose.
these normals did carry such factors.

whether ornot

The results showed that

Of the eighty—six progeny reared,

twenty—five or 29.07 per cent were bent.

This was significantly different

from the 3*3^ Per cent obtained when bent-nosed animals were outcrossed to
the normal Hoppert stock.

The difference was 25.71 * 3*55 Per cent, which

was 7.2U times its probable error.
However, the normal animals obtained from bent x bent matings were
not of the same genetic constitution as the bent-nosed animals*
tests showed this.

Progeny

The 6^*91 per cent bent-nosed, the percentage obtained

among the progeny of bent x bent matings, was significantly greater than the
29.07 per cent bent among the animals one of whose parents was a normal
from a bent x bent mating.

The difference was 35* ^ ^ 5 * ^ Per cent, which

was 6.6U times its probable error.
A part of the F

and F animals, i.e. those born at the beginning
1
2
of the genetic experiments, were put on ration XI, the high calcium ration.
The F^ animals were kept on ration II throughout the experiment, while the
F2tB were later changed to ration VII, the high phosphorus ration.

-51-

(These data are not included in table XIV.)

Of the forty-eight F

animals

which, were put on ration II, nineteen, one of which was bent, lived to 120
days of age.
bent.

Also, one of the twenty-nine which died at an earlier age was

The eighty—one F^* s which were put on ration II were transferred to

ration VII early enough so that they all survived to the 120 day age limit.
Sixteen, or 19«75 Per cent, of these F ^ s were bent.

This is a higher value

than that obtained among the F^1s reared from weaning on ration VII only,
which is not exactly consistent with the results of the nutritional experiment
which showed that ration II produced fewer bent-nosed than ration VII.
The fact that a higher percentage of bents was obtained in these
Fg's whose diet was changed from high calcium to high phosphorus than in
those fed the high phosphorus ration throughout is worth noting although the
difference between the two percentages was not quite great enough to be
significant in a statistical sense.

The difference was 9*0^ ±’3*39 P0r cent,

which was 2.67 times its probable error.

However, the difference is great

enough to indicate that here is a possible approach to the solution of the
problem.

The alteration made in the feeding increased the percentage of

bents in the

19*75 Per cent which Is not so far below 25 per cent,

that expected if the trait were controlled by a single pair of recessive
factors.

Future work on the problem could profitably be directed in search

of some possible dietary changes and the proper time for making these changes
which would increase the frequency of bent-nose.
obtained which would fit some genetic explanation.

Thus, ratios might be

-52-

Summary of Inheritance Studies
1.

Bent-nose in the rat is influenced by genetic factors.

Different types of matings produce different frequencies of bent-nose with
environmental factors constant.
2.

There is definite evidence of segregation of factors.

frequency of bent-nose in the

The

generation is significantly greater than

that in the F^ generation.
3.

It is evident that In general the factors for bent-nose are

recessive tothose for normal.

The percentage of bent animals in the F^

generation is too small to indicate any dominant factors for bent.
H.

Possibly more than one pair of genetic factors is necessary

for the manifestation of bent-nose.

Dlfhether some of these factors are

merely modifiers has not been determined, but the ratios do not Indicate
merely a single pair of factors.

LITERATURE CITED
Bethke, R. M., H. Steenbock, and Mariana T. Nelson.
Fat—soluble Vitamins.

1923.

XV Calcium and Phosphorus

Relations to Growth and Composition of Blood and
Bone with Varying Vitamin Intake.
Jour. Biol. Chem. 58 s 71-103*
Bittner, John J.

193^.

Differences Observed in an Inbred Albino
Strain of Mice Following a Change in Diet.
X.

Litter Strain.

Roscoe B. Jackson Memorial Laboratory
Nutritional Bulletin No. 1.
II.

Mortality.

Roscoe B. Jackson Memorial Laboratory
Nutritional Bulletin No. 2.
Colton, H. S.

1929*

“High brow1* Albino Rats.
Jour. Hered. 205 225—227.
Dawson, Alden B.

1927*

Further Studies on the Epiphyses of the Albino Rat
Skeleton, with Special Reference to the Vertebral
Column, Ribs, Sternum, and Girdles.
Anat. Record.

jhz 351-363*

LITERATURE CITED Continued
5*

Dawson, Alden B*

193^*

Further Studies on Epiphyseal Union in the Skeleton
of the Bat*
Anat* Record*
6. ____________ •

60: 83-86*

193^.

Additional Evidence of the Failure of Epiphyseal
Union in the Skeleton of the Bat* Studies on Wild
and Captive Gray Norway Bats*
Anat* Record.
7*

60s 5OI-5II*

Dodds, G* S* and Hazel C* Cameron*

193^*

Studies on Experimental Rickets in Bats*
Am* Jour* Anat*
8*

Geiger, Beatrice J«,

55*
H* Steenbock, and Helen T* Parsons*

La thyrism in the Bat*
Jour* of Nutrition.
9*

Gowen, John W*

6: ^27-^2*

193&*

Inheritance as it Affects Survival of Bats
Fed on a Diet Deficient in Vitamin D*
Genetics*

211 1-23*

10* Green, Charles Velmar, and Elizabeth Fekete*
Some Cranial Anomalies in Mice*
Anat* Record*

53*

187-193* •

193^*

1933*

-55LITERATORE CITED Continued
II* Green, C* V*

1932*

A Genetic Craniometric Study of Two Species of Mice
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12. Green, Harry S* N., and Wade H. Brown.

1932*

Hereditary Variations in the Skull of the Rabbit*
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1921*

Dietary Factors Influencing Calcium Assimilation*
I*

The Comparative Influence of Green and Dried

Plant Tissue, Cabbage, Orange Juice, and Cod Liver
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Jour* Biol* Chem*
lU> Heston, V. E.

US: 33~5°*

193^.

Inheritance of Bent-nose in the Rat,
(Rattus norvegicus), with Observations on the
Effect of Nutrition.
Thesis for the Degree of M* S*, Michigan State College.
15. Euestis, R* R., and Elizabeth Barto#
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27* 73-75*

1936.

-56LITERA3UBE 01 TED Continued
16. Hunt, Harrison R*, Russell Mjxter, and Dorothy Permar*
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Genetics;
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IS: 335-366*

1932.

Eight New Mutations in the Domestic Fowl*
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2: 96-97*

1935*

Pigmentation in Black-haired Bats*
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19* Kamenoff, R. S*

X935.

Development of the Flexed Tailed Mouse*
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58: 117-155*
X93I.

Studies on the Inheritance of Structural
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Am* Jour* Anat*
21* Krafka, Joseph.

US: 231-259*

1920*

qftio Effect of Temperature upon Facet Number in
the Bar—eye Mutant of Drosophila*
Jour* Gen. Physiology.

2: i409~1*6^*

22. Mixter, Bussell, and Harrison R. Hunt.

1933*

Anemia in the Flexed Tailed Mouse, Mus muscuius*
Genetics*

18* 367-367*

1933*

-57LITERATURE CITED Continued
23. Morgan, T. H.

1915 •

The Role of Environment in the Realization of a
Sex—linked Mendelian Character in Drosophila*
Am* Nat*
2b. Nadler, J. E.

385-^29*
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Effect of Temperature on Length of Vestigial Wing
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Genetics;

lit 56*4-589*

25* Orent, Elsa R*, H* D. Kruse, and E* V* McCollum*

193^*

Studies on Magnesium Deficiency in Animals*
VI*

Chemical Changes in the Rone, with Associated

Blood Changes, Resulting from Magnesium Deprivation*
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106: 573-593*

26. Reed, Sheldon C», and George D* Snell.

1931*

Harelip, a New Mutation in the House Mouse*
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51* *43-50*

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Harelip in the House Mouse*
I*

Effects of the External and Internal Environments.

Genetics.

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-58-

LITSRATURE CITED Continued
28. Eeed, Sheldon C.

1936.

Harelip in the House Mouse.
II.

Mendelian Units Concerned with Harelip and

Application of the Data to the Human Harelip Problem.
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Fluctuation in a Recessive Mendel ian Character.
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30* Sahel, Albert Edward, Julius Cohen, and Benjamin Kramer.

1935*

The Nature of the Injury to the Calcifying Mechanism
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31* __________ * 1933.
Phosphatase Activity and Calcification in Strontium Rickets.
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32. Schour, Isaac, and Margaret Cammack Smith.

193^.

The* Histological Changes in the Enamel and Dentin of the
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Univ. of Ariz. Agri. Exp. Sta. Tech. Bui. No. 52*
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1930.

Kalteschwarzung Weisser Haare und Roter Alhinoaugen.
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1919 •

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35. Sherman, H. C., and A. M. Pappenheimer.

1921.

Experimental Eickets in Eats.
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36. Sherman, H. C., and H. K. St labeling.

1929*

Quantitative Studies of Hesponses to Different Intakes
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37. ___________ •

838 **97-50^*

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38* Smith, H. V., and Margaret Cammack Smith.

1932.

Mottled Enamel in Arizona and its Correlation with the
Concentration of Eluorides in Water Supplies.
Univ. of Ariz. Agri. Exp. Sta. Tech. Bui. Ho. H3.
39* Smith, Margaret Cammack, and Edith M. Lantz.

1933*

Experimental Production of Mottled Enamel.
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-6oLITERATURE CITED Continued

1*0. Smith, Margaret CaPsaack, and Edith M. Lantz.

1933.

Changes in the Incisors of Albino Rats Accompanying a
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Pat Soluble Vitamins.
XVII.

The Induction of Growth-promoting and Calcifying

Properties in a Ration by Exposure of Ultra-violet Light.
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H2. Steenbock, H«> E. B. Hart, J. H. Jones, and A. Black.

1923*

Pat-soluble Vitamins.
XXV.

The Inorganic Phosphorus and Calcium of the Blood

Used as Criteria in the Demonstration of the Existence of
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