The electron transport system of the anaerobic Propionibacterium shermanii

1. Electron transport particles obtained from cellfree extracts of Propionibacterium shermanii by centrifugation at 105000xg for 3 hrs oxidized NADH, d,l-lactate, l-glycerol-3-phosphate and succinate with oxygen and, except for succinate, with fumarate, too.
2. Spectral investigation of the electron transport particles revealed the presence of cytochromes b, d and o, and traces of cytochrome a ₁ and a c-type cytochrome. Cytochrome b was reduced by succinate to about 50%, and by NADH, lactate or glycerol-3-phosphate to 80–90.
3. The inhibitory effects of amytal and rotenone on NADH oxidation, but not on the oxidation of the other substrates, indicated the presence of the NADH dehydrogenase complex, or “site I region”, in the electron transport system of P. shermanii.
4. NQNO inhibited substrate oxidations by oxygen and fumarate, as well as equilibration of the flavoproteins of the substrate dehydrogenases by way of menaquinone. The inhibition occurred at low concentrations of the inhibitor, and reached 80–100%, depending on the substrate tested. The site of inhibition of the respiratory activity was located between menaquinone and cytochrome b. In addition, inhibition of flavoprotein equilibration suggested that NQNO acted upon the electron transfer directed from menaquinol towards the acceptor to be reduced, either cytochrome b or the flavoproteins, which would include fumarate reductase.
5. In NQNO-inhibited particles, cytochrome b was not oxidized by oxygen-free fumarate, but readily oxidized by oxygen. It was concluded from this and the above evidence that the branching-point of the electron transport chain towards fumarate reductase was located at the menaquinone in P. shermanii. It was further concluded that all cytochromes were situated in the oxygen-linked branch of the chain, which formed a dead end of the system under anaerobic conditions.
6. Antimycin A inhibited only oxygen-linked reactions of the particles to about 50% at high concentrations of the inhibitor. Inhibitors of terminal oxidases were inactive, except for carbon monoxide.

     

Properties of a motor output system involved in the optomotor response in flies

1. The optomotor response of tethered flying houseflies (Musca domestica) has been studied at the level of the neural output which controls the activities of some non-fibrillar flight muscles (N-muscles).-a) During visually induced turning responses in a given direction some N-muscles on the right side of the thorax are synergistically active together with other N-muscles on the left side of the thorax. The same muscles are inactive during turning reactions in the opposite direction while the corresponding antagonists are now active (synopsis in Table 1).-b) The response activities of the N-mussles show a considerable variation during the course of time in spite of constant visual input.-c) There is a strong tendency for N-muscle spikes to be phase-locked with respect to the wingbeat period.-d) The findings obtained fromMusca are in accordance with the corresponding results obtained fromCalliphora (Heide, 1971b).
2. TheN-muscle activities have also been investigated in tethered flying blowflies (Calliphora erythrocephala) which tried to yaw spontaneously with both wings beating. In spontaneous left (right) turn reactions the features of the observed neural output are nearly identical with the features of the motor output showing up during visually induced left (right) turn reactions.-A different motor output pattern has been found in flies with only one wing beating.
3. The wingbeat synchronous rhythm observed in spike trains from activeN-muscles is produced in the thorax without the participation of higher stages of the fly's CNS. On the other hand no distinct rhythms can be found in spike trains fromN-muscles of non-flying flies when their motoneurons are artificially activated by non-rhythmic stimuli. Afferent information from thoracic sense organs seems to be essential for the production of the rhythm observed during flight.
4. The results about the production of the wingbeat synchronous rhythm in spike trains fromN-muscles suggest that the information derived from the motion detectors only acts to gate the output needed to achieve yaw-turn reactions. The strength of the influence of signals from the motion detectors on the output producing system can be modified by the animals “state of excitement”.
5. A model is presented which summarizes some features of information processing in the output systems supplying theN-muscles of flies. Available physiological data are discussed in relation to the model.

     

Flocculation phenomenon of Candida albicans by lysozyme

Young colonies of Sabouraud's glucose agar room temperature culture ofCandida species from human isolation were suspended in distilled water. The suspension was mixed with a solution of lysozyme and incubated in a 37° C water bath. Within 3–5 hours, various species ofCandida cells showed flocculation to varying degrees which occurred at varying periods of onset. Among sevenCandida species,Candida albicans andCandida stellatoidea showed the strongest flocculation, earliest onset and most solution clarity than did any other species.Candida stellatoidea was indistinguishable fromCandida albicans in its degree of flocculation, and in the clarity of solution.Candida species may be arranged in the following order according to their decreasing positivity in flocculation:

1. Candida albicans
2. Candida stellatoidea
3. Candida tropicalis
4. Candida krusei
5. Candida pseudotropicalis
6. Candida parapsilosis
7. Candida guilliermondii
8. Saccharomyces species may be placed afterCandida guilliermondii.

It seems possible to separate theCandida species into 3 groups by the rate of flocculation, and clarity of solution. Group I.Candida albicans andCandida stellatoidea. Group II.Candida tropicalis, C. krusei andCandida pseudotropicalis. Group III.Candida parapsilosis andCandida guilliermondii. Saccharomyces specimens (S. cerevisiae and others) were placed after group III.     

Improvements of the membrane filter method for DNA:rRNA hybridization

We describe and recommend the following improvements of DNA:rRNA membrane filter hybridization methods. One of our aims was to avoid DNA release from filter discs during hybridization.

1. Our hybridization conditions are 2 SSC in aq. dest., with 20% formamide, 50 C, overnight for 16 hr.
2. Duplexing is over in 8–10 hr.
3. Formamide has to be very pure (O.D.≤0.2/cm light path at 270 nm).
4. RNAase treatment: 250 μg/5 ml 2 SSC/filter at 37 C for 1 hr.
5. Our conditions for stepwise thermal denaturation are: 5°C steps from 50C to 90C in 1.5 SSC in 20% formamide.
6. Single-stranded DNA, fixed on membrane filters, and stored in vacuo at 4C, can be used reliably for hybridization for up to 20 months.
7. Concentrated DNA in 0.1 SSC, quick-frozen at ?50 C and stored at ?90 C for up to 2 years can be used for hybridization without much change.
8. A CsCl gradient purification step yields much purer DNA, but increases the release of DNA from filters by about 20%. Filters with 20% more DNA is a compensation.
9. rRNA can be stored for 20 months in SSC or 2 SSC at ?12C without changing the hybridization results.

     

On the possible role of respiratory activity of Acholeplasma laidlawii cells in sugar transport

1. Out of 20 exogeneous substrates only ethanol and, to a much lesser extent, lactate and pyruvate were shown to be capable of stimulating the respiration of Acholeplasma laidlawii cells. However, none of these substrates changed the initial rate of active transport of 3-O-methyl-d-glucose (3-O-MG).
2. From inhibitory analyses and spectroscopic data, it is apparent that the respiratory chain of A. laidlawii has no cytochromes and is probably not responsible for oxidative phosphorylation.
3. Valinomycin and nigericin stimulated cell respiration only in the presence of K⁺-ions, while monensin stimulated it in the presence of Na⁺-ions.
4. 3-O-MG transport was shown to be sensitive to uncouplers, ATPase inhibitors and arsenate are resistant to a majority of respiratory inhibitors tested. This suggested that there was no relationship between respiration and carbohydrate transport in the A. laidlawii cells. Further evidence was provided by the absence of respiratory stimulation during the transport of non-metabolizing carbohydrates.

     

Protease and Amylase in the digestive tract ofBarbus paludinosus

1. Protease and amylase activity in the digestive system ofBarbus paludinosus Peters (Pisces, Cyprinidae) has been investigated.
2. Chromatographic analysis showed seven amino acids to be present in both the anterior and posterior intestine. Only leucine, phenylalanine, valine, glycine and aspartic acid were positively identified.
3. In the anterior intestine chromatography revealed two sugars, but only one in the posterior intestine which was identified as glucose.
4. The pH of the intestinal fluid was found to be 5.8 and 7.8 for the fore and hind gut respectively, This correlates well with the enzyme pH optima found in in vitro experiments.
5. Protease and amylase activity was found throughout the digestive tract. Maximum proteolytic activity being present in the anterior intestine. Amylase activity is similar in both regions of the gut.
6. Correlation between the digestive enzymes and the fishes diet is briefly discussed.

     

Fumarate reduction inProteus mirabilis

1. Proteus mirabilis formed fumarate reductase under anaerobic growth conditions. The formation of this reductase was repressed under conditions of growth during which electron transport to oxygen or to nitrate is possible. In two of three tested chlorateresistant mutant strains of the wild type, fumarate reductase appeared to be affected.
2. Cytoplasmic membrane suspensions isolated from anaerobically grownP. mirabilis oxidized formate and NADH with oxygen and with fumarate, too.
3. Spectral investigation of the cytoplasmic membrane preparation revealed the presence of (probably at least two types of) cytochromeb, cytochromea ₁ and cytochromed. Cytochromeb was reduced by NADH as well as by formate to approximately 80%.
4. 2-n-Heptyl-4-hydroxyquinoline-N-oxide and antimycin A inhibited oxidation of both formate and NADH by oxygen and fumarate. Both inhibitors increased the level of the formate/oxygen steady state and the formate/fumarate steady state.
5. The site of inhibition of the respiratory activity by both HQNO and antimycin A was located at the oxidation side of cytochromeb.
6. The effect of ultraviolet-irradiation of cytoplasmic membrane suspensions on oxidation/reduction phenomena suggested that the role of menaquinone is more exclusive in the formate/fumarate pathway than in the electron transport route to oxygen.
7. Finally, the conclusion has been drawn that the preferential route for electron transport from formate and from NADH to fumarate (and to oxygen) includes cytochromeb as a directly involved carrier. A hypothetical scheme for the electron transport in anaerobically grownP. mirabilis is presented.

     

The influence of EDTA on the composition of alginate synthesized by Azotobacter vinelandii

1. During an investigation of the physiology of Azotobacter vinelandii with particular reference to polysaccharide formation, a suitable medium which was precipitate-free was developed by adding EDTA at a concentration of 50 mg/l to a basal medium containing one of eight different carbohydrates as sole carbon source.
2. Acetylated alginate was always produced by the organism when cultured under defined conditions, regardless of the carbohydrate source incorporated in the basal medium.
3. When EDTA was added to the medium, the bacteria produced acetylated polyuronides with a preponderance of mannuronic acid residues.
4. A comparison of the infrared spectra of the alginate produced by Azotobacter vinelandii and the affect of EDTA upon the mannuronic acid/guluronic acid ratios of the alginate are reported.

     

Rigorous and extended application of information theory to the afferent visual system of the cat

1. Intracellular recording were obtained from P-cells of the LGN of the cat. The impulse trains of a single presynaptic retinal ganglion cell and the postsynaptic P-cell were separated by band-pass-filtering and subsequent amplitude discrimination.
2. The rates of information and transinformation for the visual channel from the eye to a ganglion cell and to the connected P-cell were calculated. Input signals to the channel were trains of light flashes of different rate, luminance and spatial distribution.
3. Transinformation was calculated without restrictive assumptions for the code.
4. The transient behaviour of the system in response to a flash was fully considered for information calculations. Additionally, it was ensured that the state of the (adaptive) channel was considered correctly.
5. Information theory was applied in an extended way. The time courses of information transfer were calculated for various flash stimuli and compared with each other.

     

Inhibition of carotenoid synthesis in Myxococcus fulvus (Myxobacterales)

1. The inhibitory effects of CPTA, nicotine, DPA, and San 6706 on carotenogenesis in Myxococcus fulvus were investigated.
2. The effects of CPTA, D-nicotine, and L-nicotine were very similar. The action of the drugs wasadditive. The cyclization was inhibited at low doses, the introduction of the hydroxyl group at C-1′ at higher doses. Lycopene accumulated at high drug concentration. The mode of action of the inhibitors is discussed.
3. In a carotenoid mutant of M. fulvus a stimulation of the “7,8-dehydrogenase” by CPTA was observed.
4. The specific carotenoid content of bacteria was increased by DPA due to an enhanced formation of phytoene. At low doses of DPA small amounts of an intermediate carotenoid glucoside ester, a 7,8-dihydro derivative, were detected.
5. DPA was taken up by the plasma membrane. Quantitative removal of DPA by washing was not possible.
6. San 6706 specifically and reversibly blocked the desaturation of phytoene.

     

Growth requirements of blue-green algae under blue light conditions

1. Growth requirements of blue-green algae containing only the c-phycocyanin + chlorophyll a pigment system have been studied under blue light (380–540 nm) which approximates light conditions existing in subsurface waters in nature.
2. While a few species were capable of very slow photosynthetic growth on minimal medium with NO₃ ^- as nitrogen source, most species were dependent on organic compounds for comparable growth under this condition. Some organisms did quite well with only Casamino Acids as a supplement, others did well with only glucose. One species, Agmenellum quadruplicatum strain PR-6, grew only when glucose and Casamino Acids were supplied simultaneously.
3. Inhibitory effects of blue light on CO₂ fixation and nitrogen metabolism are noted as possible explanations of these responses.

     

Oxaloacetate decarboxylase from Acetobacter aceti

An oxaloacetate (OAA) decarboxylase (EC 4.1.1.3) has been purified 72-fold from Acetobacter aceti cells grown on ethanol, and its molecular weight was estimated to be about 80,000 by gel filtration. Several properties distinguished this enzyme from the OAA decarboxylase from A. xylinum:

1. It was not a constitutive enzyme; the activity was 6- to 20-fold higher in cells grown on a C₂ substrate (acetate or ethanol) than in cells grown on a C₃ compound (pyruvate or propionate).
2. The optimum pH was 7.5; a value of 5.6 was reported for the enzyme from A. xylinum.
3. The enzyme did not need a divalent cation and was not inhibited by EDTA.
4. The K _mvalue for OAA was found to be 0.22 mM. It was not affected by the addition of nicotinamide adenine dinucleotide.
5. The enzyme activity was neither inhibited by acetate nor by L-malate.

In addition, the OAA decarboxylase from A. aceti was insensitive to monovalent cations, avidin or acetyl coenzyme A.     

Biogenesis of mitochondria 20 the effects of altered membrane lipid composition on mitochondrial oxidative phosphorylation inSaccharomyces cerevisiae

1. The lipid composition of mitochondria isolated from a fatty acid desaturase mutant ofSaccharomyces cerevisiae may be extensively manipulated by growing the organism on defined supplements of unsaturated fatty acid (UFA).
2. The fatty acid composition of the mitochondrial lipids closely follows that of the whole cells from which the mitochondria are isolated. UFA-depleted mitochondria contain normal levels of sterols, neutral lipids and total phospholipids, but have much lower levels of phosphatidyl inositides.
3. UFA-depleted mitochondria possess a full complement of cytochromes, oxidase both NAD-linked and flavoprotein-linked substrates at normal rates, and have levels of succinate and malate dehydrogenases similar to those of UFA-supplemented mitochondria. However, UFA-depletion has a marked effect on the ability of cytochromec to reactivate the NADH oxidase activity of cytochromec-depleted mitochondria.
4. The efficiency of oxidative phosphorylation decreases progressively with the UFA content of the mitochondria, and oxidative phosphorylation is completely lost in mitochondria containing approximately 20% UFA.
5. The incorporation of UFA into the lipids of UFA-depleted mitochondriain vivo results in a recoupling of oxidative phosphorylation. Recoupling is insensitive to both chloramphenicol and cycloheximide, indicating that all the proteins necessary for oxidative phosphorylation are present in UFA-depleted mitochondria, and that the less of oxidative phosphorylation is a purely lipid lesion.
6. ATPase activity is apparently unaffected by UFA-depletion, but³²P_i-ATP exchange activity is lost in mitochondria which have been extensively depleted in UFA.
7. Valinomycin stimulates the respiration of UFA-supplemented mitochondria in media containing potassium, but has no effect on the respiration of UFA-depleted mitochondria, suggesting that active transport of potassium is lost as a result of UFA-depletion.

     

Life-cycles of some invertebrate taxa in a small pond together with changes in their numbers over a period of three years

1. Monthly quantitative samples of the invertebrate fauna (except Protozoa) in a small pond were taken over a period of three years. During one year, insect emergence traps were in operation. Water temperatures were recorded during the investigation.
2. The most abundant organisms in the pond were Phaenocora typhlops, Limnodrilus hoffmeisteri and Chaoborus crystallinus. Certain species of Micro-Crustacea and Chironomidae were also abundant but these groups have been dealt with elsewhere (see p. 66). Dendrocoelum lacteum, Polycelis nigra, Helobdella stagnalis, Lumbriculus aariegatus, Tubifex tubifex, Planorbis complanatus, and Asellus meridianus also occurred in considerable though lower numbers; other species occurred in low numbers.
3. The life-cycles and changes in numbers of the more numerous species are considered. The life-histories of D. lacteum, P. nigra, H. stagnalis, P. complanatus, A. meridianus are in agreement with published information. P. typhlops is seasonal in occurrence, being active from May to Sept. inclusive. Times of emergence of adults of various insect species agree with information available in the literature.
4. The life-cycle of L. hoffmeisteri in the pond is as follows: young worms hatch in spring/summer and form the bulk of the population from April to July/Aug; they mature from Aug. onwards and breeding starts in earnest from Feb./March. The life-cycle of T. tubifex is as follows: breeding starts in Feb., recruitment of young takes place from April till June, and these start to mature in Nov./Dec. It is not certain if some animals which breed in the spring/early summer survive to breed the following year.
5. The life-cycle of C. crystallinus appears to be as follows: first instars present from May to Oct., second instars from May to Dec., third instars from June till following Jan., fourth instars all the year round, pupae from May till Aug., and eggs from May to Sept. Adult emergence takes place from late April till mid-Sept.
6. A six-week drought in Oct/beginning Nov. in the second year of the investigation caused considerable mortality in most species, but most survived with only a few exceptions.

     

Investigations of carotenoids in some animals of the Adriatic Sea V. Echinodermata

The author investigated the carotenoids in the Echinodermata from Adriatic sea by means of columnar and thin-layer chromatography. The following carotenoids were identified:

- in Coscinasterias tenuispina: β-carotene, isocryptoxanthin lutein, lutein-5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin and asterinacid.

- in Marthasterias glacialis: β-carotene, echinenone, cryptoxanthin, lutein, lutein 5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin ester, astaxanthin and 3, 4-didehydro-α-carotene.

- in Paracentrotus lividus: β-carotene, echinenone, cryptothin, isocryptoxanthin, lutein, lutein-5, 6-epoxide, 4-hydroxy-4-keto-β-carotene, zeaxanthin, astaxanthin, astaxanthin ester and asterinacid.

- in Sphaerechinus granularis: ,β-carotene, echinenone, cryptoxanthin, lutein, lutein-5, 6-epoxide, astaxanthin and guaraxanthin.

     

Comparative studies of carotenoids in the fauna of the Gullmar Fjord (Bohuslan,Sweden) III. Echinodermata

The author investigated the presence of various carotenoids in the Echinodermata from Gullmar Fjord (Bohuslan, Sweden) by means of columnar and thin-layer chromatography. The investigations revealed the presence of the following:

- inHenricia sanguinolenta:β-carotene, echinenone, canthaxanthin, guraxanthin, lutein-5, 6-epoxide and astaxanthin.

- inAmphiura filiformis: canthaxanthin, cryptoxanthin, lutein, lutein-5, 6-epoxide, isozeaxanthin, zeaxanthin, astaxanthin and 4-hydroxy-4-keto-β-carotene.

- inAmphipholis squamata:β-carotene, cryptoxanthin, lutein, lutein-5, 6-epoxide, astaxanthin, astaxanthin ester, asterin-acid and rubixanthin derivative.

- inOphiopholis aculeata: canthaxanthin, cryptoxanthin, isozeaxanthin, astaxanthin, astaxanthin ester, asterinacid, 4-hydroxy-4-keto-β-carotene, hydroxy rubixanthin and gazaniaxanthin-like substances.

- inOphiothrix fragilis: canthaxanthin, lutein-5, 6-epoxide, isozeaxanthin, astaxanthin, astaxanthin ester, 4-hydroxy-4-keto-β-carotene, and hydroxy rubixanthin.

- inAntedon petatus:canthaxanthin, guaraxanthin, isozeaxan-thin, zeaxanthin, astaxanthin, astaxanthin ester and 4-keto-4-ethoxy-β-carotene.

- inEchinocardium cordatum:β-carotene,γ-carotene, canthaxanthin, lutein, isozeaxanthin, zeaxanthin, astaxanthin and astaxanthin ester.

- inSpatangus purpureus: isozeaxanthin, astaxanthin, astaxanthin ester and 4-hydroxy-4-keto-β-carotene.

     

Time course of the Houseflies' landing response

The landing response of stationary flying houseflies Musca domestica has been recorded on video tape. The leg movements were quantitatively evaluated. It could be demonstrated that:

1. only the first two pairs of legs are involved in the reaction (Fig. 1). Prothoracic tarsi are lifted beyond the head, mesothoracic tarsi are lowered and moved sidewards (Fig. 2a and b).
2. the movement of the tarsal tips is mainly due to an opening of one single joint per leg, i.e. the femurtibia joint of the prothoracic leg (Fig. 2c), and the coxa-femur joint of the mesothoracic leg.
3. the landing reaction is a fixed action pattern which does not seem to require further sensory input once it is released (Fig. 4d).
4. the landing responses to a light-off stimulus and to expanding patterns with different angular velocities are indistinguishable (compare Fig. 3a-c with Fig. 2a-c). The only parameter that is obviously dependent on the stimulus conditions, is the latency of the reaction (Fig. 4a-c).

     

Context-dependent stuttering

Bei einer Untersuchung des Stotterns in der deutschen Umgangssprache machten wir folgende Beobachtungen:

1. Das gestotterte Phonem wird häufig von einem identischen Phonem begleitet (definiert als das ?induzierende Phonem“) welches sowohl vor wie nach dem gestotterten Phonem auftreten kann.
2. Gewöhnlich folgte das induzierende Phonem dem gestotterten Phonem.
3. Der Abstand zwischen induzierendem und gestottertem Phonem war geringer als bei Zufälligkeit zu erwarten.
4. Induzierende und gestotterte Phoneme befanden sich gewöhnlich in identischen Silbenpositionen.
5. Gestotterte Phoneme traten in der Regel bei betonten Silben auf.

Um diese Beobachtungen zu erklären erschienen uns drei Annahmen erforderlich:

1. Sprach-Output ist hierarchisch bestimmt. Silben und Phoneme sind Glieder in dieser Hierarchie.
2. Unterschwellige Erregbarkeit ist in dieser Hierarchie stärker bei betonten als bei unbetonten motorischen Programmen.
3. Ähnliche Programme (sowohl auf Silbenals auch Phonemniveau) inhibieren einander.

Diese Annahme gibt zugleich eine mögliche Erklärung für Blockierung und Längung — Phänomene, die ebenfalls in der Sprache von Stotterern auftreten. Unsere Beobachtungen bieten also eine mögliche Lösung für das Rätsel des Stotterns.     

ATP-driven succinate oxidation in the catabolism of Desulfuromonas acetoxidans

The oxidation of succinate with elemental sulphur in Desulfuromonas acetoxidans was investigated using a membrane preparation of this bacterium. The following results were obtained:

1. The preparation catalyzed the oxidation of succinate with sulphur and NAD. These reactions were dependent on ATP and were abolished by the presence of protonophores or dicyclohexylcarbodiimide (DCCD).
2. The membrane preparation also catalyzed the reduction of fumarate with H₂S or with NADH. These activities were not dependent on ATP and were not affected by protonophores or DCCD.
3. By extraction-reincorporation experiments it could be shown that menaquinone is involved in electron transport between H₂S and fumarate and between NADH and fumarate.
4. The membrane fraction catalyzed the reduction of the water-soluble menaquinone-analogue dimethylnaphthoquinone (DMN) by succinate, H₂S, or NADH, and the oxidation of DMNH₂ by fumarate. These activities were not dependent on the presence of menaquinone and were not influenced by ATP.
5. The activities involving succinate oxidation or fumarate reduction were similarly sensitive to 2(n-nonyl)-4-hydroxyquinoline-N-oxide, while H₂S and NADH oxidation by DMN were not affected by the inhibitor.

It is concluded that the catabolism of D. acetoxidans involves the energy-driven oxidation of succinate with elemental sulphur or NAD as electron acceptors and that menaquinone is a component of the electron transport chain catalyzing these reactions.