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A K Fok

Publications and source records attributed to A K Fok.

At least 37 records · Page 2Linked to original sources
PubMed1989

Intracellular binding of wheat germ agglutinin by Golgi complexes, phagosomes, and lysosomes of Paramecium multimicronucleatum.

The compartments of the Paramecium digestive system were investigated with wheat germ agglutinin (WGA). By use of cryosectioning or Lowicryl K4M embedding combined with pulse-chase studies and WGA-gold labeling, WGA binding sites were located on membranes of the phagosome-lysosome system, including all four stages of digestive vacuoles, the discoidal vesicles, acidosomes, and lysosomes. In addition, the contents of lysosomes, cisternae at the trans face of Golgi stacks, and coated and uncoated blebs and vesicles at the putative trans Golgi network bind to WGA. Crystal-containing vacuoles characteristic of mid-log to stationary-phase cultures are enclosed by heavily labeled membranes. Alveoli underlying the plasma membrane sometimes contain binding sites, particularly on their outer membranes. Ciliary membranes previously shown to be labeled with WGA-FITC are negative in frozen thin and Lowicryl K4M sections. The presence of WGA binding sites on the trans face of the Golgi stack is the first indication in ciliated protozoa, such as Paramecium, of probable Golgi complex involvement in glycosylation similar to that in higher organisms. WGA-labeled coated vesicles in the endoplasm apparently lose their coats and coalesce to form lysosomes. Our study shows that WGA can be used as a specific intracellular marker of all digestive system membranes and of lysosomal content. These results support and extend our published scheme of membrane flow and recycling in Paramecium by providing another means of demonstrating membrane relationships.

Animals↗
PubMed1988

A survey of lectin binding in Paramecium.

To better understand the general distribution of glycoproteins and the distribution of specific glycoprotein-bound sugar residues in Paramecium, a survey of the binding pattern of selected lectins was carried out in P. tetraurelia, P. caudatum, and P. multimicronucleatum. Lectins studied were concanavalin A (Con A), Griffonia simplicifolia agglutinins I and II (GS I and GS II), wheat germ agglutinin (WGA), Ulex europaeus (UEA I), peanut agglutinin (PNA), Ricinis communis toxin (RCA60) and agglutinin (RCA120), soybean agglutinin (SBA), Bauhinia purpurea agglutinin (BPA), Dolichos biflorus agglutinin (DBA), and Maclura pomifera agglutinin (MPA). Those giving the most distinctive patterns were Con A, GS II, WGA, UEA I, and PNA. No significant differences were found between the three species. Concanavalin A, a mannose/glucose-binding lectin, diffusely labeled the cell surface and cytoplasm and, unexpectedly, the nuclear envelopes. Events of nuclear division, and nuclear size and number were thus revealed. Both WGA and GS II, which are N-acetylglucosamine-binding lectins, labeled trichocyst tips, the cell surface, and the oral region, revealing stages of stomatogenesis. The lectin WGA, in addition, labeled the compartments of the phagosome-lysosome system. The lectin PNA, an N-acetyl galactosamine/galactose-binding protein, was very specific for digestive vacuoles. Finally, UEA I, a fucose-binding lectin, brightly labeled trichocysts, both their tips and body outlines. We conclude that a judicious choice of lectins can be used to localize glycoproteins and specific sugar residues as well as to study certain events of nuclear division, cellular morphogenesis, trichocyst discharge, and events in the digestive cycle of Paramecium.

Animals↗
PubMed1988

Phagosome formation in Paramecium: effects of solid particles.

Digestive vacuole (DV) formation in Paramecium can be separated into four steps: sequestration and recycling of the spent DV membrane, sweeping and concentrating of particles into the nascent DV, vacuole growth, and the release of the nascent DV. How the size, load and surface charge of solid particles affected formation and subsequently the digestive processes in log-phase cells were investigated. Pulsing cells continuously with beads resulted in an initial linear increase, followed by a steady state, of labelled DVs. Above a certain threshold concentration, the rate of formation and the size of the DVs formed (as well as the steady state) all increased with increasing bead load, so that for a 16-fold increase in bead load, a corresponding fourfold increase in the incorporation of the recycled membrane into the DVs was observed. The threshold values, which depended on the sensitivity of the technique used to score these DVs, were lowered as bead size increased. The steady state of labelled DVs was shown to correspond to a strict balance between the formation and defecation rates as shown by efflux measurements and pulse-chase protocols using two consecutive labels. The duration of the pulse required to reach these steady states was inversely proportional to the logarithm of the bead number at low bead loads but remained constant at higher bead loads. The formation rates and the DV size were similar when cells were pulsed with beads from 0.5 to 3 microns, but DV size increased using beads of 4.5 and 5.7 microns. The maximal bead size that could be ingested was approximately 10 microns.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Characterization of monoclonal antibodies to trichocyst antigens in Paramecium.

Ten mouse monoclonal antibodies (mAbs) were raised against trichocyst contaminants present in crude or enriched lysosome fractions of Paramecium multimicronucleatum. Using an indirect immunofluorescence assay (IFA) and immunogold labelling on frozen thin sections, epitopes were located on the outer edge, cortex and the core of the trichocyst body, as well as the sheath covering the tip. Except for the two on the tip, epitopes were reactive after SDS-PAGE under non-reducing conditions. Four mAbs (131C1E8, A1-3, A16-2, D7) were directed to a trio of bands of 37, 34 and 29 (x 10(3] Mr from the beaded or meshlike trichocyst body sheath. A fifth mAb (135B9E7), directed to epitopes on the cortex inside the beaded body sheath, reacted strongly with the 37 and 34 bands, but weakly with the 29 x 10(3) Mr band. The last three mAbs (270D5, 22D7F2, D8) were reactive with one or more of three families of antigens found on the trichocyst core. mAb 270D5 reacted mainly with the 34 and 29 (x 10(3] Mr bands of the family containing the above trio, while mAb 22C7F2 reacted consistently with the 47 x 10(3) band of the higher Mr family but variably with both the trio of bands and the 17 x 10(3) band of the lower Mr family. mAb D8, which was directed to epitopes on the trichocyst core and small vesicles in the endoplasm, reacted only with the 29 x 10(3) Mr band. The mAbs were cross-reactive with the trichocysts of P. primaurelia, P. tetraurelia, P. caudatum and P. calkinsi with some small variation in blotting patterns.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Ionophores and weak bases inhibit phagolysosomal proteolysis in Paramecium.

In a recent report we showed that ionophores and weak bases inhibit digestive vacuoles (DV) acidification primarily and lysosome-DV fusion secondarily but have no effect on lysosome-DV fusion when acidification is normal. In this study we attempted 1) to show that fluorescein isothiocyanate (FITC)-albumin taken up by phagocytosis could be used for a sensitive proteolytic assay, 2) to use this assay to determine the effect of ionophores and weak bases on proteolysis and 3) to learn how an inhibition of acidification and/or lysosome-DV fusion would affect proteolysis. When cells were pulsed with FITC-albumin and latex beads for 3 min and chased, the amount of albumin degraded increased linearly from 9 to 27 min, reaching a plateau by 30 min, and was inhibited by leupeptin and pepstatin A by 47 to 89%. These results showed that the degradation of FITC-albumin occurred in the phagolysosomes. When added before acidification had commenced, carbonyl cyanide p-trifluoro-methoxyphenyl hydrazone (FCCP), monensin and NH4Cl partially inhibited lysosome-DV fusion (25-50%) and strongly inhibited proteolysis by 64 to 79%. Added between acidification and lysosome-DV fusion, fusion was unaffected while proteolysis was reduced by 40 to 50%. Added after lysosome-DV fusion was completed, proteolysis was still reduced by the same amount. Chloroquine at 0.25 mM had no effect on proteolysis except when added before acidification, it inhibited fusion by 22% and proteolysis by 16%. These data, together with those published recently, showed that 1) ionophores and weak bases inhibited acidification first, lysosome-DV fusion second and proteolysis third, but they also inhibited proteolysis directly and independent of the prior steps and 2) the proteolysis inhibitory effects were additive.

Animals↗
PubMed1987

Phagosomal acidification in Paramecium: effects on lysosomal fusion.

Phagosomes of paramecia and amoeba and endosomes of fibroblasts and other mammalian cells are acidified prior to lysosomal fusion. The question, whether the phagosomal acidification process in paramecia is required for phagosome-lysosome fusion, was studied using ionophores, weak bases, and cytochalasin B (CB) in combination with monoclonal antibodies, acid phosphatase (AcPase) cytochemistry, and lysosome morphometry. Digestive vacuoles (DVs) of known ages were treated and examined. In untreated cells, lysosome binding to the membrane of the acidified DV increased linearly with age and reached a maximum before lysosome-DV fusion. When the fusion of the acidosomes with the very young DVs was prevented by CB, causing a block in the normal vacuole-pH drop, lysosome binding to the DVs as well as the rate and extent of lysosome-DV fusion were all greatly reduced. These effects of CB were reversible. When present prior to acidification, three ionophores and two weak bases did not inhibit the acidosome-DV fusion but raised the phagosomal pH and reduced both the rates of DV acidification and of lysosome-DV fusion. However, when added after acidification but prior to lysosome-DV fusion, five of the six perturbants studied did not inhibit this fusion but prolonged the period when DVs remained AcPase positive. Lastly, lysosome-DV fusion rates were found to be related to acidification rates. We conclude that an inhibition in acidosome-DV fusion or a reduction in both the acidification rate and vacuolar-pH drop would inhibit lysosome-DV fusion.

Acid Phosphatase↗
PubMed1985

Processing of digestive vacuoles in Tetrahymena and the effects of dichloroisoproterenol.

The digestive-lysosomal system in Tetrahymena has been extensively studied; however, the various vacuole stages and the existence of a required processing period prior to defecation have not been clearly defined. In this study the presence of such a required processing period and the rate of DV defecation in Tetrahymena thermophila were determined. Like the cycle in Paramecium, a digestive cycle in Tetrahymena consisted of two periods: the processing period was 45 min and the defecation period was approximately 2 h, making the complete cycle approximately 3 h. During the defecation period vacuole egestion followed the kinetics of a first-order rate reaction and had a rate constant of 0.0187/min and a t1/2 of 37 min (82 min into the cycle). Using the naphthol AS-TR phosphate-hexazotized rosanilin method to visualize acid phosphatase activity at the light microscopic level, DVs became positive beginning at 10 min. The number of positive DVs increased to a maximum of 13% when DVs were 20-min old and declined to 5-7% beyond 30 min. Although dichloroisoproterenol (DCI) has been reported by others to stimulate vacuole defecation, we found it inhibited the defecation rate. The extent of inhibition depended on the age of the DVs when exposed to DCI. Vacuole formation was completely blocked in cells preexposed to 40 microM DCI for only 10 min; however, upon further exposure, cells could recover from this inhibition. The time required for complete recovery increased with increasing DCI concentrations. If DCI was given to cells simultaneously with latex beads, it was found to exert a dose-dependent inhibitory effect on DV formation.(ABSTRACT TRUNCATED AT 250 WORDS)

Acid Phosphatase↗
PubMed1985

Modulation of the digestive lysosomal system in Paramecium caudatum. II. Physiological effects of cytochalasin B, colchicine and trifluoperazine.

The heterophagic pathway of the digestive lysosomal system in Paramecium caudatum includes at least four steps: digestive vacuole (DV) formation, acidification-condensation, lysosomal fusion-digestion, and defecation. The second and the third require about 20 min, during which DVs are not egested. Because these steps occur at predictable intervals, the mechanism for each can be explored by exposing labeled DVs to different drugs prior to each step. In this study the effects of cytochalasin B (CB), colchicine and the calmodulin antagonist, trifluoperazine (TFP), were studied. All three drugs inhibited DV formation in a dose-dependent manner when cells were pulsed with latex beads and a drug simultaneously. TFP was cytotoxic above 5 microM. Vacuole formation was completely shut down when cells were pre-exposed to 5 microM TFP for 13 min. At this level, the duration of the acidification step was lengthened, and the rate of defecation decreased with increasing exposure. These results suggest that these inhibitory effects may be more related to TFP's cytotoxicity than to its action on calmodulin-mediated process. Colchicine at 1 mM had no effect on the third or fourth step, but inhibited the acidification step so that DVs were egested later and at a slower rate. Exerting a differential effect on all four steps, CB inhibited DV release from the cytopharynx, egestion of defecation-competent DVs at the cytoproct and lengthened the duration but did not block the lysosomal fusion-digestion step of the acidic DVs; it was most potent in blocking acidification, which prevented both lysosomal fusion with the labeled DVs as well as DV egestion, the latter for more than 50 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Modulation of the digestive lysosomal system in Paramecium caudatum. III. Morphological effects of cytochalasin B.

Morphological explanations for the effects of cytochalasin B (CB) on the digestive vacuole system of Paramecium caudatum were sought using electron-microscopic studies. Cytochalasin B (0.3 mM) essentially stopped vacuole release without stopping vacuole growth. Acidosome fusion with the new vacuoles was inhibited and was accompanied by the formation of discoidal vesicle-lined clumps of microfilament-like material next to these vacuoles. Vacuole membrane retrieval at the open cytoproct was inhibited and was accompanied by the loss of the microfilament network typical of this region. At 0.15 mM CB vacuole release and fusion of the acidosomes and lysosomes with the vacuoles were not stopped, but membrane retrieval at the cytoproct was affected. Cytochalasin B at the higher concentration also affected the movement of the horseradish peroxidase (HRP)-labeled vesicles away from the nascent vacuole. Membrane retrieval by the formation of tubules of 35 to 50 nm diameter from membrane of early and late vacuole stages was also affected by CB. It is conceivable that all of these effects were the result of the disruption of membrane associated actin networks. By contrast, several highly organized microfilamentous systems in the cortex and oral region of these cells appeared to be unaffected by CB or dimethyl sulfoxide (DMSO) at the concentrations used. These results show that CB-susceptible structures assume a key role in the normal functioning of the digestive vacuole system and in vacuole membrane retrieval and reutilization in Paramecium.

Animals↗
PubMed1985

Rapid and sensitive assays for phagosomal acidification in Paramecium and Tetrahymena.

Biochemical and cytochemical procedures were developed to measure the rate of phagosomal acidification for phagosomal pH ranging from 5 to 2.5. These assays were based on the pH-dependent inactivation with time of horseradish peroxidase (HRP) activity, a result attributable to the dissociation of this enzyme to a colorless protein and ferriprotoporphyrin in acidic solutions. When preincubated in buffers of varying pH, the rate of HRP inactivation followed a sigmoid curve, with the highest rate of inactivation between 4.3 and 3.5 when using citrate-phosphate buffer and between pH 3.4 and 2.8 when using the universal ABC buffer. This inactivation was temperature but not concentration dependent. When Paramecium caudatum, members of the P. aurelia complex or Tetrahymena thermophila was pulsed briefly with HRP and small fluorescent beads, the loss of HRP activity, measured biochemically in cell homogenates and/or cytochemically in phagosomes, was rapid and followed the kinetics of a first-order rate reaction. Both assays gave similar values for the rate constant for acidification and similar rates of inhibition when P. caudatum was exposed to a proton ionophore, carbonyl cyanide p-trifluoromethoxyphenyl hydrazone. These assays can readily be adapted to other phagocytic cells as long as a rapid procedure is available for removing all unphagocytosed HRP and latex beads. These procedures are sensitive and rapid thus allowing many samples to be quickly prepared and analyzed.

Animals↗
PubMed1984

Retrieval of lysosomal membrane and acid phosphatase from phagolysosomes of Paramecium caudatum.

Little is known about the fate of lysosomal membrane in phagocytic cells. Because the age of the digestive vacuoles in Paramecium caudatum can be easily determined, we have been able to study the dynamic membrane events in the older vacuoles. Late in the phagolysosomal stage (DV-III) the vacuole membrane undergoes a burst of tubule formation. The tubules expand into vesicles which have characteristics resembling lysosomes in both thin sections and freeze-fracture replicas. The tubules also contain acid phosphatase activity when they arise from acid phosphatase-reactive vacuoles. We conclude that after active digestion lysosomal membrane is retrieved in whole or in part along with some membrane-associated hydrolases. A logical extension of these results is that the lysosome-like vesicles, after being recharged with hydrolases by fusing with primary lysosomes, are recycled back to DV-II for reuse.

Acid Phosphatase↗
PubMed1984

Modulation of the digestive-lysosomal system in Paramecium caudatum. I. Effects of temperature.

The heterophagic pathway of the digestive-lysosomal system in axenically grown Paramecium caudatum is divisible into vacuole formation, vacuole acidification-condensation, lysosomal fusion-digestion and defecation. These four processes can be separated in time, thus permitting the study of the effects of temperature on each process. The optimal growth temperature for this cell was 27 degrees C. The rate of digestive vacuole (DV) formation at varying temperatures was represented by a skewed bell-shaped curve having an optimum between 28 and 30 degrees C. The time course for the acidification-condensation step was lengthened below 26 degrees C, but was not accelerated above this temperature. The rate but not the extent of vacuole condensation was decreased at 19 and 22 degrees C. Temperature increase above 22 degrees C shortened, slightly, the duration of the lysosomal fusion-digestion process, whereas below 22 degrees C small temperature decreases greatly extended this period. Within a given experiment the rates of defecation were proportional to temperatures above 17 degrees C. However, these rates varied widely among different experiments. Interestingly, the activation energies for both the formation and defecation processes averaged 19 kcal/mol. Furthermore, Paramecium appeared to readily adapt to environmental temperature changes, since the length of the processing periods and the rates of defecation were similar in cells with or without a 24 h acclimation. These results indicated that the four processes in the digestive cycle in P. caudatum are distinct but each is energy-dependent.

Animals↗
PubMed1983

Nonlysosomal vesicles (acidosomes) are involved in phagosome acidification in Paramecium.

Although acidification of phagocytic vacuoles has received a broadened interest with the development of pH-sensitive fluorescent probes to follow the pH changes of vacuoles and acidic vesicles in living cells, the mechanism responsible for the acidification of such vacuoles still remains in doubt. In previous studies of the digestive vacuole system in the ciliate Paramecium caudatum we observed and described a unique population of apparently nonlysosomal vesicles that quickly fused with the newly released vacuole before the vacuole became acid and before lysosomes fused with the vacuole. In this paper we report the following: (a) these vesicles, named acidosomes, are devoid of acid phosphatase; (b) these vesicles accumulate neutral red as well as acridine orange, two observations that demonstrate their acid content; (c) cytochalasin B given 15 s after exposure of the cells to indicator dye-stained yeast will inhibit the acidification of yeast-containing vacuoles; and that (d) we observed using electron microscopy, that fusion of acidosomes with the vacuole is inhibited by cytochalasin B. We conclude that the mechanism for acidification of phagocytic vacuoles in Paramecium resides, at least partially if not entirely, in the acidosomes.

Animals↗
PubMed1983

Effects of dimethylsulfoxide (DMSO) on the digestive-lysosomal system in Paramecium caudatum.

The effects of DMSO (dimethylsulfoxide) on cell growth and on the digestive-lysosomal system of axenically grown Paramecium caudatum were studied. A general protocol of exposing cells to different concentrations of DMSO at the beginning of each of the four processes in the digestive cycle enabled us to analyze the effect of DMSO at each step. Vacuole formation and the beginning of a digestive cycle were initiated by adding latex beads to the cells. Maximum cell densities at stationary phase of growth were found to be inversely proportional to DMSO between 0.5 and 1.75%, and the duration of the generation time was exponentially proportional. At 2% DMSO cellular division was completely blocked, and above 2% it was cytotoxic. P. caudatum survived for 8 h in 4% DMSO and died instantaneously in 10%. This inhibitory effect on growth was reversible, though this reversibility might depend on the duration and level of DMSO exposure. DMSO exerted a dose- and time-dependent inhibitory effect on the rate of DV formation but had little effect on the acidification-condensation and the lysosome fusion-digestion processes. The size of the DV formed was also reduced, and this effect was dose-but not time-dependent; vacuole size reduction occurred immediately with DMSO exposure, and no further reduction was observed during exposures of up to 24 h. DMSO at 3 and 4% inhibited vacuole defecation, but the cells could overcome this inhibition when exposed to DMSO for longer periods.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Phagosome fusion vesicles of paramecium. I. Thin-section morphology.

Ultrastructural studies of the digestive system of Paramecium caudatum focusing on the first 5 min of digestive-vacuole age reveal a set of vesicles, named phagosome fusion vesicles (PFVs), which fuse with the digestive vacuole just after the vacuoles are released from the cytopharynx and concomitant with vacuole acidification. Serial thin-sections of vacuoles labeled with horseradish peroxidase (HRP) and/or latex beads in pulse-chase studies were observed. PFVs, irregularly shaped, electron-translucent vesicles ranging from a small diameter to over 1 micro, are first seen in the region of the cytopharynx where they bind to the nascent vacuole membrane. Within 30 sec of vacuole release the PFVs fuse with the vacuole where they remain for a brief time connected to the vacuole by a narrow annulus. HRP-reaction product is found in vacuoles but not in PFVs before PFVs fuse with the vacuoles. After fusion with PFVs HRP is quickly inactivated. Tubular extensions of vacuole membrane then form between the fused PFVs. By 3 to 5 min both PFVs and tubules disappear from the vacuole surface and lysosomes appear in their place. We believe the tubules are pinched off as PFV membrane is being added to the vacuole. Microfilaments coat the membrane during all these dynamic events. Since the pH of the vacuole becomes acid during the first few minutes, we are now looking for a direct correlation between PFV fusion and acidification.

Animals↗
PubMed1983

Phagosome fusion vesicles of Paramecium. II. Freeze-fracture evidence for membrane replacement.

Phagosome fusion vesicles (PFVs), a new population of relatively large granules in Paramecium caudatum which fuse with the first stage of digestive vacuoles (DV-I) shortly after these vacuoles are released from the cytopharynx (their site of formation), have been studied by using the freeze-fracture technique. Identification of PFVs is possible in the resulting replicas at all sites where they are commonly found in thin sections, at the cytopharynx, bound but not fused with nascent digestive vacuoles and fused with released vacuoles in the cell's posterior end. These PFVs have membranes which do not resemble the membranes of the forming digestive vacuole membrane or the discoidal vesicle membranes from which vacuole membrane is derived. Their smooth E-fracture face with only 50 to 100 intramembrane particles (IMPs) per micrometers 2 and particulate P-face (approximately 2500 IMPs/micrometers) do resemble the second vacuole stage (DV-II) which is characterized by a smaller diameter and acid pH. Evidence is presented for PFV fusion with the DV-I and for membrane replacement, at least in part, as the DV-I becomes a DV-II. Membrane replacement entails first adding PFVs to the DV-I and then removing the original discoidal vesicle-derived membrane as tubules as the vacuole condenses. Implications of the possible role of PFVs in forming intravacuolar symbiotic relationships are also discussed.

Animals↗
PubMed1981

Axenic Paramecium caudatum. III. Biochemical and physiological changes with culture age.

As Paramecium caudatum passes through the lag, log and stationary phases of the culture cycle, cellular protein content, polar (PL) and neutral (NL) lipid contents, marker enzyme activities, rate of digestive vacuole formation and cellular viability undergo characteristic changes. Maximal protein content (60 ng/cell) and enzyme activities ranging from 7 nmoles for catalase to 0.2 pmoles/cell/min for alkaline phosphatase were observed between days 2 and 4. This active metabolism paralleled the fine structure of 1 to 3 day-old cells which contained extensive foci of rough endoplasmic reticulum (RER) partially bordered by Golgi stacks and the rapid depletion of those lipid fields accumulated during day 1. Decrease in protein content and enzyme activities in late log phase indicated a slowing of cellular synthesis. The lipids in the medium were largely depleted and accounted for the low lipid uptake of 14 ng/cell on day 5 as compared with 615 ng/cell on day 1. Yet a vast amount of protein lysate was still available in the culture medium. During stationary phase, catalase activity remained constant, but activities of alkaline and acid phosphatases and 5'nucleotidase declined gradually to low levels, while those of Ca2+-ATPase and malate dehydrogenase declined precipitously. Only 25% of the maximal activities of the latter two enzymes remained by the end of stationary phase. A ten-fold increase in the cellular PL and NL content was already observed 24 h postinoculation. This accumulation was used for subsequent growth and cell divisions; PL declined exponentially and NL less steeply between days 1 and 6. PL remained level (5 ng/cell) throughout stationary phase while NL declined further to 1 ng/cell by day 11. The rate of digestive vacuole formation was constant (6.3 +/- 0.5 DV/5 min pulse) during the entire log phase, then declined from 4.4 on day 6 to 0.22 on day 11. When early to mid-stationary-phase cells were subcultured, some lag in growth was seen; a definite lag was observed when inoculating with late-stationary-phase cells. When early-death-phase cells were given fresh nutrients, many died; the surviving ones became fully rejuvenated after 48 h. The biochemical and physiological data from this study are correlated with the morphological study of the companion paper.

Alkaline Phosphatase↗