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A new, cheap, high-power micromanipulator
Notes and Brief Articles
493
survey of twenty isolates the roughest' smooth walled' spore found was that of T. vinosa (Hypocrea vinosa Cooke) illustrated in PI. 29, fig. 6. The warts on this spore have dimensions approximately one-sixth of those of T. viride in PI. 29, fig. 2 and would thus support the concept of discontinuity. However, the present investigation has covered a limited number of isolates, and in particular only two isolates of T. viride, and a wider survey would be required before reaching any conclusions. The technique described in this paper has been successfully applied to six species of Penicillium. The spores of a seventh species showed regular and rapid collapse. It is suggested that the technique outlined here may prove applicable to the spores of genera other than Trichoderma. I wish to thank Professor J. Webster for the gift of a number of cultures used in this work, and Dr D. L. Hawksworth for assistance in reclassifying certain IMI cultures of Trichoderma. I also wish to acknowledge the help of Mrs C. Cummings in preparing the electron micrographs. REFERENCES
RIFAI, N. A. & WEBSTER,]. (1966). Culture studies on Hypocrea and Trichoderma. II. H. aurea-viridis and H. rufa f. sterilis f.nov, Trans. Br. mycol. Soc. 49, 289-296. RIFAI, M. A. (1969). A revision of the genus Trichoderma. Mycol. Pap. No. rr6. EXPLANATION OF PLATE
29
Spores of Trichoderma Fig. Fig. Fig. Fig. Fig. Fig.
I.
2. 3. 4. 5. 6.
T. viride SHD-M 261 I. Phialospores x 7500. T. viride IMI 45553. Phialospores x 7500. T. koningii WEe 25277A. Phialospores x 7500. T. koningii WEe 25277A. Part of phialospore X30000. T. pseudokoningii IMI 92027 (listed as T. viride). Part of phialospore x 30000. T. oinosa WEe 4589. Phialospores x 7500.
A NEW, CHEAP, HIGH-POWER MICROMANIPULATOR R. F. O. KEMP
Department of Botany, Unioersity of Edinburgh Amicromanipulator* is described which can be constructed very cheaply since it is made of 'Perspex' acrylic sheet and only the halving and tapping of four phosphor-bronze balls involves precision engineering. It has no massive base and there is full coincidence of movements of main control with the needle when viewed under the microscope. It can be adjusted to give zero lateral movement or be set for use as a microdissector, and is ideal for the quick isolation offungal spores and other work on agar surfaces. The two types of micromanipulator in most general use are the pneu'" Registered design. Trans. Br. mycol. Soc. 55 (3), (1970). Printed in Great Britain
494
Transactions British Mycological Society
matic model of de Fonbrune (1937) and the simpler ball-lever model of Barer & Saunders-Singer (1948). The former is well suited for microsurgery and injection and the latter for the isolation of fungal spores and manipulation of hyphae growing on an agar surface. Both of these models are somewhat expensive if they are to be used relatively infrequently. The model described below is based on the one made by Saunders-Singer but o
o
o
o
0
-f-- 1
I I I I I
0
I I I
0
0
0
!
:
I
I
2
9a
10 a
I
6
I3 \7
IT--~rSt=::l[::::========:jt=::::;:::t=::;i::::;:::;td:~ 7 a 12
14
13
·9
10
Fig. I. Detailed construction of the instrument. The numbered parts are (I) base plate, (2) top plate, (3) T-piece, (4) T-piece plate, (5) T-piece plate supports, (6) T-piece pivot, (7) ball bearings, (7a) groove, (8) T-piece spring, (g) main control, (ga) main control hemispheres, (10) coarse control, (lOa) coarse control hemispheres, (II) retaining rods, springs and balls, (12) needle column, (13) clamp, (14) hammer.
with several important modifications. It is made of 6 mm black' Perspex ' and its structure may be seen in Fig. I. It is cheap enough to be used in equipping student laboratories. There are three main differences between the new model and that of Saunders-Singer. First, vertical movement of the needle is obtained by having two phosphor-bronze hemispheres (9a) at the top of the main control (9). The top hemisphere is fixed but the lower one is tapped and moves on a short length of thread. The hemispheres rest in countersunk holes and although both are free to slide in their cups the lower one does not Trans. Br. mycol. Soc. 55 (3), (Ig70). Printed in Great Britain
Notes and Brief Articles
495
rotate when the main control is rotated. The anticlockwise rotation of the main control raises the top plate (2) and thus the needle. This arrangement eliminates the need for any sort of base. When the hemispheres are together no lateral movement of the needle is possible if the main control is moved laterally. Anticlockwise rotation of the main control separates the hemispheres so that lateral movement of the needle is possible at any level of reduction. With the hemispheres less than I mm apart the instrument can be used as a high-power micromanipulator. When they are more than 5 mm apart it becomes a microdissector. The second difference is the method of obtaining the coarse movement of the needle in the vertical plane. Instead of having this adjustment working through a base it is obtained by having the two ball bearings (7) supporting the top plate mounted on a pivotted T -piece (3). Anticlockwise rotation of the coarse control raises the needle as the T -piece with its ball bearings tilts down. A spring (8) mounted below the T-piece pushes it upwards thus lowering the needle when the coarse control is turned in a clockwise direction. This control also has two hemispheres one of which is tapped. The third important difference is that the main control is located between the ball bearing supports and the needle. This results in all movements of the main control being identical to those of the needle when viewed under the microscope. The Saunders-Singer model has the ball bearing supports between the main control and the needle, resulting in a reversal of movements when the control is moved away from the operator. With these modifications it is possible to dispense with a massive base and to have only two moving ' Perspex ' components (2, 3). These, together with the base plate (I) are easily made. Of the small number of metal components only the halving and tapping of four phosphor-bronze balls need be done with precision. The ball bearing support (7) which is not running in the groove has to be fixed in a slightly deeper hole than the other to keep the top plate level. The balls on the retaining rods are also made of phosphor-bronze. The top one is fixed and the other is drilled so that it can slide on the rod. The tension of the spring is controlled by a small adjustable clamp. The T-piece pivot rod (6) can be inserted into one of two holes to give two settings of the coarse adjustment. For use the instrument has to be mounted on a rigid support such as a large weighted block of wood. Being such a simple instrument no provision is made for the mechanical centering of the needle in the microscope field. This can be done either by moving the whole instrument or by moving the needle and clamp on the needle column. When the needle is focused in the centre of the field the instrument can be set to work at the required level of reduction, by rotating the main control. The needle can be kept in focus while this is done by counteracting the movement of the main control by adjusting the coarse control. The anticlockwise rotation of both controls raises the needle. If the instrument is set for use as a microdissector it may be necessary to slacken off the springs on the retaining rods. Just above the handle of the main control is a small spring-mounted hammer Trans. Br, m)'col. Soc. 55 (3), (1970). Printed in Great Britain
496
Transactions British Mycological Society
which can be used for knocking fungal spores out of a microloop. This is necessary as the main control handle cannot be tapped against the control rod as in the Saunders-Singer model. It is essential that the lower cups of both controls are kept free of grease otherwise the lower hemisphere will rotate. If necessary the cups can be cleaned by hand using a countersink. Clearly the new instrument does not have the same degree of rigidity as those with massive bases but this has proved no handicap in the isolation of single fungal spores or bacteria on an agar surface. The very slight amount of bending which is possible when the main control is pulled down towards the bench can be used to lower the needle the last few microns on to the agar surface without having to rotate the control. The vertical movement of the main control involves only a slight lateral movement of the needle as it is roughly 30 ern from the two ball bearings which act as the fulcrum. It is a great deal less than with the de Fonbrune model which operates in a distinct arc. The de Fonbrune is also much slower to operate because of the resistance of its pneumatic system. In comparison with the Saunders-Singer model much less practice is needed in dissecting with two instruments as the movements of needle and control are identical. Instead of having two manipulators for fixing microelectrodes a double model can be made which has a larger base plate so that two top plates and sets of controls can be mounted in parallel. Different left-hand and right-hand models are unnecessary as the baseplates can be made identical on both sides so that the other parts can be assembled on either side. REFERENCES
BARER, R. & SAUNDERS-SINGER, A. E. (1948). A new single-control micromanipulator. 0...]1 microsc. Sci. Sg, 439-447. DE FONBRUNE, P. (1937). Micromanipulateur pneumatique et microforge. Paris: Societe Industrielle d'Imprimerie.
NEW NAMES FOR TWO WEST AFRICAN RUST FUNGI F. C. DEIGHTON
Commonwealth Mycological Institute, Keto In accordance with Art. 59 of the International Code of botanical Nomenclature 1966, new names are needed for the following two West African species of Uredinales. Ravenelia albiziae-zygiae Deighton, nom.nov.
Syn. Ravenelia z:;ygiae H. Sydow, Annls mycol. 36, 157, 1938 based on telia, not R. z:;ygiae H. Sydow, Annis mycol. 35, 248, 1937 based on uredia. The name R. z:;ygiae Syd. (1937) was based on two syntypes from Sierra Leone and was validly published but illegitimate since it was published in a genus characterized by the perfect state though only the imperfect (uredial) state was described. The fact that part of one of the syntype Trans. Br. mycol. Soc. 55 (3), (1970). Printed in Great Britain
493
survey of twenty isolates the roughest' smooth walled' spore found was that of T. vinosa (Hypocrea vinosa Cooke) illustrated in PI. 29, fig. 6. The warts on this spore have dimensions approximately one-sixth of those of T. viride in PI. 29, fig. 2 and would thus support the concept of discontinuity. However, the present investigation has covered a limited number of isolates, and in particular only two isolates of T. viride, and a wider survey would be required before reaching any conclusions. The technique described in this paper has been successfully applied to six species of Penicillium. The spores of a seventh species showed regular and rapid collapse. It is suggested that the technique outlined here may prove applicable to the spores of genera other than Trichoderma. I wish to thank Professor J. Webster for the gift of a number of cultures used in this work, and Dr D. L. Hawksworth for assistance in reclassifying certain IMI cultures of Trichoderma. I also wish to acknowledge the help of Mrs C. Cummings in preparing the electron micrographs. REFERENCES
RIFAI, N. A. & WEBSTER,]. (1966). Culture studies on Hypocrea and Trichoderma. II. H. aurea-viridis and H. rufa f. sterilis f.nov, Trans. Br. mycol. Soc. 49, 289-296. RIFAI, M. A. (1969). A revision of the genus Trichoderma. Mycol. Pap. No. rr6. EXPLANATION OF PLATE
29
Spores of Trichoderma Fig. Fig. Fig. Fig. Fig. Fig.
I.
2. 3. 4. 5. 6.
T. viride SHD-M 261 I. Phialospores x 7500. T. viride IMI 45553. Phialospores x 7500. T. koningii WEe 25277A. Phialospores x 7500. T. koningii WEe 25277A. Part of phialospore X30000. T. pseudokoningii IMI 92027 (listed as T. viride). Part of phialospore x 30000. T. oinosa WEe 4589. Phialospores x 7500.
A NEW, CHEAP, HIGH-POWER MICROMANIPULATOR R. F. O. KEMP
Department of Botany, Unioersity of Edinburgh Amicromanipulator* is described which can be constructed very cheaply since it is made of 'Perspex' acrylic sheet and only the halving and tapping of four phosphor-bronze balls involves precision engineering. It has no massive base and there is full coincidence of movements of main control with the needle when viewed under the microscope. It can be adjusted to give zero lateral movement or be set for use as a microdissector, and is ideal for the quick isolation offungal spores and other work on agar surfaces. The two types of micromanipulator in most general use are the pneu'" Registered design. Trans. Br. mycol. Soc. 55 (3), (1970). Printed in Great Britain
494
Transactions British Mycological Society
matic model of de Fonbrune (1937) and the simpler ball-lever model of Barer & Saunders-Singer (1948). The former is well suited for microsurgery and injection and the latter for the isolation of fungal spores and manipulation of hyphae growing on an agar surface. Both of these models are somewhat expensive if they are to be used relatively infrequently. The model described below is based on the one made by Saunders-Singer but o
o
o
o
0
-f-- 1
I I I I I
0
I I I
0
0
0
!
:
I
I
2
9a
10 a
I
6
I3 \7
IT--~rSt=::l[::::========:jt=::::;:::t=::;i::::;:::;td:~ 7 a 12
14
13
·9
10
Fig. I. Detailed construction of the instrument. The numbered parts are (I) base plate, (2) top plate, (3) T-piece, (4) T-piece plate, (5) T-piece plate supports, (6) T-piece pivot, (7) ball bearings, (7a) groove, (8) T-piece spring, (g) main control, (ga) main control hemispheres, (10) coarse control, (lOa) coarse control hemispheres, (II) retaining rods, springs and balls, (12) needle column, (13) clamp, (14) hammer.
with several important modifications. It is made of 6 mm black' Perspex ' and its structure may be seen in Fig. I. It is cheap enough to be used in equipping student laboratories. There are three main differences between the new model and that of Saunders-Singer. First, vertical movement of the needle is obtained by having two phosphor-bronze hemispheres (9a) at the top of the main control (9). The top hemisphere is fixed but the lower one is tapped and moves on a short length of thread. The hemispheres rest in countersunk holes and although both are free to slide in their cups the lower one does not Trans. Br. mycol. Soc. 55 (3), (Ig70). Printed in Great Britain
Notes and Brief Articles
495
rotate when the main control is rotated. The anticlockwise rotation of the main control raises the top plate (2) and thus the needle. This arrangement eliminates the need for any sort of base. When the hemispheres are together no lateral movement of the needle is possible if the main control is moved laterally. Anticlockwise rotation of the main control separates the hemispheres so that lateral movement of the needle is possible at any level of reduction. With the hemispheres less than I mm apart the instrument can be used as a high-power micromanipulator. When they are more than 5 mm apart it becomes a microdissector. The second difference is the method of obtaining the coarse movement of the needle in the vertical plane. Instead of having this adjustment working through a base it is obtained by having the two ball bearings (7) supporting the top plate mounted on a pivotted T -piece (3). Anticlockwise rotation of the coarse control raises the needle as the T -piece with its ball bearings tilts down. A spring (8) mounted below the T-piece pushes it upwards thus lowering the needle when the coarse control is turned in a clockwise direction. This control also has two hemispheres one of which is tapped. The third important difference is that the main control is located between the ball bearing supports and the needle. This results in all movements of the main control being identical to those of the needle when viewed under the microscope. The Saunders-Singer model has the ball bearing supports between the main control and the needle, resulting in a reversal of movements when the control is moved away from the operator. With these modifications it is possible to dispense with a massive base and to have only two moving ' Perspex ' components (2, 3). These, together with the base plate (I) are easily made. Of the small number of metal components only the halving and tapping of four phosphor-bronze balls need be done with precision. The ball bearing support (7) which is not running in the groove has to be fixed in a slightly deeper hole than the other to keep the top plate level. The balls on the retaining rods are also made of phosphor-bronze. The top one is fixed and the other is drilled so that it can slide on the rod. The tension of the spring is controlled by a small adjustable clamp. The T-piece pivot rod (6) can be inserted into one of two holes to give two settings of the coarse adjustment. For use the instrument has to be mounted on a rigid support such as a large weighted block of wood. Being such a simple instrument no provision is made for the mechanical centering of the needle in the microscope field. This can be done either by moving the whole instrument or by moving the needle and clamp on the needle column. When the needle is focused in the centre of the field the instrument can be set to work at the required level of reduction, by rotating the main control. The needle can be kept in focus while this is done by counteracting the movement of the main control by adjusting the coarse control. The anticlockwise rotation of both controls raises the needle. If the instrument is set for use as a microdissector it may be necessary to slacken off the springs on the retaining rods. Just above the handle of the main control is a small spring-mounted hammer Trans. Br, m)'col. Soc. 55 (3), (1970). Printed in Great Britain
496
Transactions British Mycological Society
which can be used for knocking fungal spores out of a microloop. This is necessary as the main control handle cannot be tapped against the control rod as in the Saunders-Singer model. It is essential that the lower cups of both controls are kept free of grease otherwise the lower hemisphere will rotate. If necessary the cups can be cleaned by hand using a countersink. Clearly the new instrument does not have the same degree of rigidity as those with massive bases but this has proved no handicap in the isolation of single fungal spores or bacteria on an agar surface. The very slight amount of bending which is possible when the main control is pulled down towards the bench can be used to lower the needle the last few microns on to the agar surface without having to rotate the control. The vertical movement of the main control involves only a slight lateral movement of the needle as it is roughly 30 ern from the two ball bearings which act as the fulcrum. It is a great deal less than with the de Fonbrune model which operates in a distinct arc. The de Fonbrune is also much slower to operate because of the resistance of its pneumatic system. In comparison with the Saunders-Singer model much less practice is needed in dissecting with two instruments as the movements of needle and control are identical. Instead of having two manipulators for fixing microelectrodes a double model can be made which has a larger base plate so that two top plates and sets of controls can be mounted in parallel. Different left-hand and right-hand models are unnecessary as the baseplates can be made identical on both sides so that the other parts can be assembled on either side. REFERENCES
BARER, R. & SAUNDERS-SINGER, A. E. (1948). A new single-control micromanipulator. 0...]1 microsc. Sci. Sg, 439-447. DE FONBRUNE, P. (1937). Micromanipulateur pneumatique et microforge. Paris: Societe Industrielle d'Imprimerie.
NEW NAMES FOR TWO WEST AFRICAN RUST FUNGI F. C. DEIGHTON
Commonwealth Mycological Institute, Keto In accordance with Art. 59 of the International Code of botanical Nomenclature 1966, new names are needed for the following two West African species of Uredinales. Ravenelia albiziae-zygiae Deighton, nom.nov.
Syn. Ravenelia z:;ygiae H. Sydow, Annls mycol. 36, 157, 1938 based on telia, not R. z:;ygiae H. Sydow, Annis mycol. 35, 248, 1937 based on uredia. The name R. z:;ygiae Syd. (1937) was based on two syntypes from Sierra Leone and was validly published but illegitimate since it was published in a genus characterized by the perfect state though only the imperfect (uredial) state was described. The fact that part of one of the syntype Trans. Br. mycol. Soc. 55 (3), (1970). Printed in Great Britain