Improvements relating to the Inoculation of Soil for the Cultivation of Leguminous Plants.
Year: 1895

page_text: Date of Application, 12th June, 1895
Complete Specification Left, 9th Mar., 1896-Accepted, 25th Apr., 1896
PROVISIONAL SPECIFICATION.
Improvements relating to the Inoculation of Soil for the Cultivation
of Leguminous Plants.
We, Dr. FRIEDRICH NOBBE, Privy Counsellor and Professor, and Dr. LORENZ
HILTNER, Chemist, of Tharand, in the German Empire, do hereby declare the
mature of this invention to be as follows:-
Since the function of the root nodules or tubercles of the Legumino æ in the
supply of nitrogen to these plants has been discovered by the fundamental
researches of Hellriegel, we have been working on this problem for a number of
years (since 1889), and have more especially examined the bacteria identified in
the said root nodules or tubercles in the meantime by Von Beyerinck and obtained
by him in pure cultures, by agricultural researches, in order to determine the
relation between these bacteria and the reception of the free inactive nitrogen of
the air in the soil by the various kinds of Leguminosa.
These researches have resulted in the first place, in the confirmation of the, at that
time, still disputed fact that the introduction of these bacteria into soil, produces,
without exception, in soil previously free from these bacteria, the root nodules or
tubercles on the plants in question having papilionaceous flowers and enables these
growths to effect the assimilation of the free nitrogen. A soil inoculated with
these bacteria, even when it contains absolutely no nitrogen in an assimilable form,
so that the plants without any such inoculation would starve, enables the
Leguminosa to produce as rich a yield of dry material and nitrogen as they would
otherwise produce only if grown in a richly manured soil containing much assimilable
nitrogen.
It has been established by us, as an entirely new fact, that the tubercle bacteria
of the various Papilionacea are of full strength (i.e. in the production of nodules or
tubercles) only with that species from whose root tubercles they were themselves
obtained, with nearly allied species they are of weaker strength, and with
systematically different species they are useless. Bacteria cultures from pea-roots,
for example, are quite useless for Robinia plants, whilst they promote the growth
of peas in an extremely energetic manner, and the allied vetches somewhat more
feebly; and inversely, the bacteria from Robinia nodules or tubercles are quite
efficient with Robinia plants, in a lesser degree with Colutea, and are absolutely
useless with peas.
At first sight it might possibly be thought that the production, transport and
distribution of such large masses of crude inoculating material as would appear to
be necessary for the sufficient impregnation or treatment of large areas of land,
would be very difficult and costly, and therefore not practicable, whilst there wouldi
also be the danger that, in the crude inoculating material, besides the active
bacteria of the root nodules or tubercles, there would be carried from field to field
at the same time microscopic organisins which would be detrimental to growth and
would more or less interfere with the action of the inoculating material.
Our process is, however, free from any such objections as those above mentioned,
inasmuch as bacteria bred in quantities directly from the nodules or tubercles of
the Leguminosa in pure cultures, are used as the inoculating material. Farmers
are thereby placed in a position to make land which was unfruitful by reason of its
lack of nitrogen, fit for the cultivation of fodder and other plants belonging to the

page_text: order of the Leguminosa, and to ensure and increase the yield of better soils. This
inoculation has, moreover, an essentially practical bearing in connection with the
so called "green manuring."
Our process of inoculating land with tubercle bacteria is to be carried out as
follows:-
The active bacteria for growth with the Leguminose are delivered to the farmer
in glass tubes which contain pure colonies thereof in Agar-gelatine having suitable
additions, such, for instance, as sugar, asparagine, or an aqueous extract of the
green substance of Leguminosa. According to circumstances the bacteria can also
be prepared for transport in fluid cultures. The colonies in Agar-gelatine are
distributed in water together with the Agar-gelatine by the user, after removing
the stopper, the contents of one glass tube being added to from 1 to 3 litres of
water which is previously mixed with a suitable material, such, for instance, as in
aqueous extract of the green substance of Leguminosa, sugar, or asparagine, for
propagating the bacteria, this material being delivered therewith. Preferably the
glass tube is laid in the water until the Agar-gelatine is dissolved.
Immediately before sowing, the whole of the emulsion prepared as above
mentioned is poured over the seeds. The amount of water added for each kind of
seed is so proportioned that, after the seeds have been thoroughly and uniformly
moistened by a careful working over by hand, a surplus of liquid will still remain.
For clover seed, for example, for 20 kilogrammes of seed, the admixture of 3 litres
of water with the contents of 3 glasses (cach glass having, for instance, a capacity
of about 30 cubic centimetres) of inoculating material is sufficient. For more bulky
seeds a somewhat larger amount of water is required. A sufficient quantity of
sand or earth from the field to be sown is then gradually added with careful stirring
until the mass of seeds is in a suitable condition for sowing by hand or by means of
a sowing machine.
Instead of inoculating the seed, a small quantity of sand (or earth) can be
saturated with the emulsion containing the bacteria and carefully mixed. After
being dried in the air, this impregnated mass of earth is distributed and worked
into the soil to a depth of about 10 centimetres.
Dated 11th day of June 1895.
D. YOUNG & Co.,
11 & 12 Southampton Buildings, London, W.C.,
Agents for the Applicants.
Improvements relating to the Inoculation of Soil for the Cultivation
of Leguminous Plants.
We, Dr. FRIEDRICH NOBBE, Privy Counsellor and Professor, and Dr. LORENZ
HILTNER, Chemist, of Tharand, in the German Empire, do hereby declare the
nature of this invention and in what manner the same is to be performed, to be
particularly described and ascertained in and by the following statement :-
Since the function of the root nodules or tubercles of the Leguminosa in the
supply of nitrogen to these plants has been discovered by the fundamental researches
of Hellriegel, we have been working on this problem for a number of years
(since 1889) and have more especially examined the bacteria identified in the said
root nodules or tubercles in the meantime by Von Beyerinck and obtained by him
in pure cultures in his agricultural researches, in order to determine the relation
between these bacteria and the reception of the free inactive nitrogen of the air
in the soil by the various kinds of Leguminosa.
These researches bave resulted in the first place, in the confirmation of the,

page_text: at that time, still disputed fact that the introduction of these bacteria into soil,
produces, without exception, in soil previously free from these bacteria, the root
nodules or tubercles on the plants in question having papilionaceous flowers, and
enables these growths to assimilate the free nitrogen. A soil inoculated with these
bacteria, even when it contains absolutely no nitrogen in an assimilable form, so
that the plants without any such inoculation would starve, enables the Leguminosa
to produce as rich a yield of dry material and nitrogen as they would otherwise
produce only if grown in a richly manured soil containing much assimilable
nitrogen.
It has been established by us, as an entirely new fact, that the tubercle bacteria
of the various Papilionace are of full strength (i.e. in the production of nodules
or tubercles) only with that species from whose root tubercles they were them-
selves obtained, with nearly allied species they are of weaker strength, and with
systematically different species they are useless. Bacteria cultures from pea-roots,
for example, are quite useless for Robinia plants, whilst they promote the growth
of peas in an extremely energetic manner, and the allied vetches somewhat more
feebly; and inversely, the bacteria from Robinia nodules or tubercles are quite
efficient with Robinia plants, in a lesser degree with Colutea, and are absolutely
useless with peas.
At first sight it might possibly he thought that the production, transport and
distribution of such large masses of crude inoculating material, as would appear
to be necessary for the sufficient impregnation or treatment of large areas of land,
would be very difficult and costly, and therefore not practicable, whilst there
would also be the danger that in the crude inoculating material, besides the active
bacteria of the root nodules or tubercles, there would be carried from field to field
at the same time microscopic organisms which would be detrimental to growth and
would more or less interfere with the action of the inoculating material.
Our process is, however, free from any such objections as those above mentioned,
inasmuch as bacteria bred in quantities directly from the nodules or tubercles of
the Leguminosa, in pure cultures, are used as the inoculating material. Farmers
are thereby placed in a position to make land which was unfruitful by reason of
its lack of nitrogen, fit for the cultivation of fodder and other plants belonging to
the order of the Leguminosa, and to insure and increase the yield of better soils.
This inoculation has, moreover, an essentially practical bearing in connection with
the so-called "green manuring."
Our process of inoculating land with tubercle bacteria is to be carried out as
follows:-
The active bacteria for growth with the Leguminosa are delivered to the farmer
in glass tubes which contain pure colonies therof in Agar-gelatine having suitable
additions, such as sugar, asparagine, or an aqueous extract of the green substance
of Leguminosa. According to circumstances the bacteria can also be prepared for
transport in fluid cultures. The colonies in Agar-gelatine are distributed in water
together with the Agar-gelatine by the user after removing the stopper, the contents
of one glass tube to from 1 to 3 litres of water which is previously mixed with a
suitable material, such as an aqueous extract of the green substance of the
Leguminosa, sugar, or asparagine, for propagating the bacteria, this material being
delivered therewith. Preferably the glass tube is laid in the water until the
Agar-gelatine is dissolved.
Immediately before sowing, the whole of the emulsion prepared as above
mentioned is poured over the seeds. The amount of water added for each kind
of seed is so proportioned that, after the seeds have been thoroughly and uniformly
moistened by a careful working over by hand, a surplus of liquid will still remain.
For clover seed, for example, for 20 kilogrammes of seed, the admixture of 3 litres
of water with the contents of 3 glasses (each glass having for instance a capacity
of about 30 cubic centimeters) of inoculating material is sufficient. For more
bulky seeds a somewhat larger amount of water is required. A sufficient quantity
of sand or earth from the field to be sown is then gradually added with careful

page_text: stirring until the body of seed is in a suitable condition for sowing by hand or by
means of a sowing machine.
Instead of inoculating the seed, a small quantity of sand (or earth) can be
saturated with the emulsion containing the bacteria and carefully mixed. After
being dried in the air, this impregnated mass of earth is distributed and worked
into the soil to a depth of about 10 centimeters.
Having now particularly described and ascertained the nature of our said inven-
tion and in what manner the same is to be performed, we declare that what we
claim is:-
1. The employment of a pure culture of the bacteria of the root nodules or
tubercles of the Leguminosae for the purpose of promoting the growth of such
plants, substantially as hereinbefore described.
2. The herein described method or process of inoculating soil, for the culture of
Leguminosae, with pure cultures of the bacteria of the root nodules or tubercles of
the same or a closely allied species of Leguminosa, by mixing pure-culture tubercle-
bacteria either with the seeds together with fine granular material, such as sand,
earth or the like, and a food substance for the bacteria, or with the latter materials
alone, and then distributing the mixture over the soil.
3. A tube or bottle containing pure colonies of tubercle bacteria of the
Leguminosa in Agar-gelatine together with a suitable substance for propagating
such bacteria, substantially as described.
Dated this Oth day of March 1896
D. YOUNG & Co.,
11 & 12 Southampton Buildings, London, W.C.,
Agents for the Applicants.