Microbiological analysis of rock cod (Sebastes spp.) stored under elevated carbon dioxide atmospheres

The numbers and types of microorganisms on fresh rock cod fillets and fillets stored in air or in a modified atmosphere (MA; 80% CO(2), 20% air) at 4 degrees C were compared. Samples were analyzed after 0, 7, 14, and 21 days of storage. The isolation plates were incubated aerobically, anaerobically, or under MA at 4, 20, or 35 degrees C. After 7 days of storage in air, the fillets were obviously spoiled and had a 3- to 4-log cycle increase in microbial counts. Plate counts increased more slowly on MA-stored fillets. After 21 days, the counts on the latter had increased only 2 log cycles, and the fillets did not seem spoiled. The microbial flora changed greatly during MA storage. Only Lactobacillus spp. (70%) and an Aeromonas sp.-like isolate (30%) were found on plates incubated aerobically at 4 and 20 degrees C, and only Lactobacillus spp. was found on plates incubated aerobically and anaerobically at 35 and at 20 degrees C under MA. Isolation plates incubated at 20 degrees C in air gave the highest counts in the shortest incubation time and the greatest diversity of bacterial types recovered. No Vibrio parahaemolyticus, Staphylococcus aureus, or Clostridium botulinum type E were isolated from the fresh or MA-stored fillets.     

Effect of pH and Oxygen Tension on Staphylococcal Growth and Enterotoxin Formation in Fermented Sausage

Commercial fermented 0sausages that contained significant numbers of viable coagulase-positive staphylococci were found to have the growth localized in the outermost areas of the sausage where oxygen tension was highest. Staphylococci were found to be more acid-tolerant aerobically than anaerobically. With chemical acidulation of sausage, growth could be controlled both aerobically and anaerobically with approximately 1.5% glucono delta lactone. Biological acidulation with a high inoculum of Pediococcus cerevisiae inhibited anaerobic staphylococcal growth but failed to suppress aerobic growth completely. A staphylococcal count of approximately 4 × 10⁷ cells/g of sausage appeared to be necessary to produce detectable enterotoxin A within 24 hr in sausage. A minor difference existed in the relative rates of production of the different types of enterotoxin. Detectable enterotoxin A was produced in 24 hr in sausage held in atmospheres containing 10, 15, and 20% oxygen. In an atmosphere containing 5% oxygen, toxin was detected after 48 hr of incubation. No toxin was detected after 120 hr under anaerobic conditions. Most staphylococcal strains tested initiated growth and produced detectable enterotoxin aerobically at a pH of 5.1 in broth media. Anaerobically, however, most strains failed to produce detectable enterotoxin below pH 5.7.     

Sampling and storage conditions of rainbow trout liver affects monooxygenase and conjugation enzymes

1. The effect of storage conditions of rainbow trout (Salmo gairdneri) liver on monooxygenase and conjugation enzyme activities was studied. Fish livers or whole fish were frozen and stored for various periods of time at -4, -20 or -80 degrees C. 2. Freezing the whole fish at -20 degrees C affected the biotransformation enzyme activities dramatically. The loss of monooxygenase activity exceeded up to one-tenth of the initial rate in 17 days. UDP-Glucuronosyltransferase activity increased 50%. Glutathione S-transferase appeared to be the most durable enzyme. 3. When the whole fish were stored in an ice-bath at -4 degrees C for up to 24 hr the activities measured decreased only half of that when frozen for 3 days. 4. When it is impossible to freeze the tissues studied in liquid nitrogen the activities are best preserved when whole, decapitated, bled fish are kept in an ice-bath for less than 24 hr.     

Comparison of cold storage and perfusion of dog livers on function of tissue slices

We compared how two methods of hypothermic preservation affect physiological functions of tissue slices of dog liver. Livers were preserved by either (i) cold storage (CS) in Collins' solution or (ii) continuous perfusion (P) with a perfusate, containing hydroxyethyl starch, sodium gluconate, adenosine, and potassium phosphate, recently developed in our laboratory. Livers were cold stored for 6 to 8, 24, or 48 hr, and perfused for 24 or 72 hr. Tissue slices of preserved livers were incubated at 30 degrees C and analyzed for volume control, electrolyte-pump activity (K and Na), and adenine nucleotide concentration. Also, mitochondria were isolated after preservation to quantify respiratory activity. Slice functions of livers preserved for short periods (6 to 8 hr by CS and 24 hr by P) were similar to those for control livers. After normothermic incubation, the mean (+/- SD) water content of tissue (expressed per unit dry mass of tissue) was 2.3 +/- 0.3 kg/kg for control, 2.6 +/- 0.4 kg/kg for 6- to 8-hr CS, and 2.5 +/- 0.5 kg/kg for 24-hr P. Longer periods of preservation resulted in cell swelling, and water content was 3.3 +/- 0.4 kg/kg for 24- to 48-hr CS and 2.8 +/- 0.3 kg/kg for 72-hr P. The mean (+/- SD) K/Na ratio was nearly normal for livers preserved for short periods: 3.7 +/- 0.5 for control, 4.1 +/- 0.2 for 6- to 8-hr CS, and 3.3 +/- 0.4 for 24-hr P.(ABSTRACT TRUNCATED AT 250 WORDS)     

Relationship between dead cells and DNA fragmentation in bovine embryos produced in vitro and stored at 4 degrees C.

DNA fragmentation and its relationship with dead cells were examined in bovine blastocysts produced in vitro and stored at 4 degrees C for 1-5 days. Survival and development to the hatching and hatched blastocyst stage decreased with increasing storage time. Both were significantly lower at 72 hr than at 48 hr. None of the embryos stored for 120 hr developed to the hatching or hatched blastocyst stage. The proportion of dead cells per embryo increased progressively as the time of storage increased, until 69% of embryonic cells were dead after 120 hr of storage. There was no significant difference between the proportions of DNA fragmentation per embryo stored for 0 and 24 hr (12% vs 16%). However, the proportion of DNA fragmentation in embryos stored for longer than 48 hr was significantly greater than that in embryos stored for less than 24 hr. There were no significant differences among those stored for longer than 48 hr (28-33%). These results suggest that the reduced developmental competence of bovine embryos stored at 4 degrees C is characterized by necrotic change rather than apoptotic change.     

Effect of temperature and ripening stages on membrane integrity of fresh-cut tomatoes

The shelf-life of fresh-cut tomatoes mainly depends on loss of tissue integrity and firmness that occurs also in intact fruits after long-term cold storage due to chilling injury. Round-fruit tomatoes (Solanum lycopersicum L.) cv. Jama were stored in 1.1-L plastic (polyethylene) fresh-cut produce containers as 10.0-mm-thick tomato slices and as intact tomatoes at 4 ± 0.5 °C. The aim of this work was to study the loss of membrane integrity and biochemical processes involved in membrane disruption. Electrolyte leakage and lipid peroxidation were studied at different stages of maturity: mature green, pink (PK), fully ripe and two different storage temperatures: 4 and 15 °C. The tomato slices of PK stage stored at 4 °C did not show changes for both parameters, while significant increase in membrane leakage and lipid peroxidation was observed at 15 °C, especially after 24 h of storage. The enzymes showed a simultaneous increase in their activities with a rise in electrolyte leakage and lipid peroxidation after 7 days of storage. Finally, phospholipase C (PLC) and phospholipase D (PLD) were investigated for intact fruit and tomato slices stored at 4 °C. The PLC had higher activity compared with PLD. In conclusion, the loss of membrane integrity in fresh-cut tomatoes is mainly affected by ripening stages, storage temperature and duration. The wounds enhance the PLC and PLD activities and they play a role late during storage.     

Circadian oxygen consumption rhythm of the flour beetle, Tribolium confusum

Oxygen consumption is circadian (about 24 hr) rhythmic in sterilized Tribolium confusum adults kept on a 12 hr light-12 hr dark regimen. High values for oxygen consumption occur shortly after ‘light-on’ and low ones at the beginning of the dark span. When the light-dark cycle is advanced by 6 hr, the oxygen consumption rhythm is phase-shifted within about 6 days; with a delay of the light-dark schedule (a return to the initial regimen) the oxygen consumption rhythm re-adjusts in 3 days. This directedness or polarity of the phase-shift adjustment constitutes a quantitative circadian system characteristic shared by organisms studied thus far.     

Poly(ethylene glycol)-conjugation and deoxygenation enable long-term preservation of hemoglobin-vesicles as oxygen carriers in a liquid state

The stability of hemoglobin vesicles (HbV) as an oxygen infusion was tested during the storage for 1 year at 4, 23, and 40 degrees C. The surface of the HbV was modified with poly(ethylene glycol) (PEG), and the suspension was deoxygenated with nitrogen bubbling. The samples stored at 4 and 23 degrees C showed a stable dispersion state for 1 year, though the sample stored at 40 degrees C showed the precipitation and decomposition of vesicular components, a decrease in pH, and 4% leakage of total Hb after 1 year. The PEG chains on the vesicular surface stabilize the dispersion state and prevent the aggregation and fusion due to their steric hindrance. The original metHb content (ca. 3%) before the preservation gradually decreased to less than 1% in all the samples after 1 month due to the presence of homocysteine inside the vesicles which consumed the residual oxygen and gradually reduced the trace amount of metHb. The rate of metHb formation was strongly dependent on the partial pressure of oxygen, and no increase in metHb formation was observed due to the intrinsic stability of the deoxygenated Hb. Preservation at 4 and 23 degrees C slightly reduced P(50) (increased the oxygen affinity) from 38 Torr to 32 and 31 Torr, respectively. These results indicate the possibility that HbV suspension can be stored at room temperature for at least 1 year.     

Development of a cold storage solution for pancreas preservation

Canine pancreas tissue slices were incubated at 5 degrees C for 24 hr in solutions containing different saccharides (raffinose, sucrose, mannitol, or glucose). At the end of incubation tissue water (TW expressed as kg H2O/kg dry wt) was determined as a measure of tissue edema. Tissue edema was greatest in slices stored in Eurocollins (EC) solution (TW = 4.96 +/- 0.14) which contains glucose for osmotic pressure. The degree of edema was decreased by saccharides in proportion to their molecular mass: mannitol (MW = 180, TW = 3.84 +/- 0.08), sucrose (MW = 348, TW = 3.54 +/- 0.08), and raffinose (MW = 594, TW = 3.30 +/- 0.07). Tissue edema was also greatest in slices incubated in solutions containing the smallest molecular mass anions: Cl- (TW = 4.02 +/- 0.16), gluconate (TW = 3.69 +/- 0.10), and lactobionate (TW = 3.28 +/- 0.13). Cold storage of the intact pancreas in EC solution for 24 hr did not induce as much edema as in slices (TW = 2.88 +/- 0.10). However, on isolated reperfusion at normothermia (37 degrees C) the pancreas became edematous (TW = 3.33 +/- 0.12). Storage of the pancreas in a lactobionate-raffinose solution did not induce edema after 90 min of normothermic reperfusion. The suppression of tissue edema in the pancreas may be essential to obtaining long-term preservation (24-72 hr) of this organ which is currently limited to about 6-8 hr in EC solution. The newly developed lactobionate-raffinose solution appears to control tissue edema in both tissue slices and the intact-flushed out organ.     

Live rats resulting from injection of oocytes with spermatozoa freeze-dried and stored for one year

This study was designed to examine whether rat spermatozoa after freeze-drying and 1-year storage can participate in full-term development following intracytoplasmic sperm injection (ICSI). Cauda epididymal spermatozoa from Crlj:Wistar rats were frozen in liquid nitrogen (LN(2)), first dried for 14 hr at 0.37 hPa and then for 3 hr at 0.001 hPa. The dried spermatozoa were stored for 1 year in a desiccator at +25 degrees C, or in a refrigerator at +4 degrees C, or in LN(2) at -196 degrees C. Controls consisted of sperm that had only been frozen and stored in LN(2). After being stored, spermatozoa were sonicated to dissociate the sperm tail and were injected into oocytes from superovulated Slc:SD rats. The respective fertilization rates of oocytes injected with frozen sperm, or with freeze-dried sperm stored at +25, +4, and -196 degrees C were 79%, 75%, 70%, and 73%. However, the corresponding cleavage rates of injected oocytes were 63%, 1%, 38%, and 36%. After transfer of >80 zygotes of each group into recipients, the respective percentages of full-term normal offspring resulting from frozen sperm or from freeze-dried sperm stored at +25, +4, and -196 degrees C were 36%, 0%, 7%, and 14%. These results demonstrate that the storage temperature significantly influenced the likelihood of term development of rats produced by injection of oocytes with freeze-dried spermatozoa. Chromosomal analysis of the rat spermatozoa in the ICSI oocytes indicated that chromosomal aberration in freeze-dried spermatozoa stored at +25 degrees C (100%) occurred more frequently than in frozen control spermatozoa (41%) and freeze-dried spermatozoa stored at -196 degrees C (35%), and the frequency of chromosomal aberrations in freeze-dried spermatozoa stored at +4 degrees C (65%) was the intermediate. In conclusion, rat spermatozoa freeze-dried and stored at +4 degrees C for 1 year are capable of participating in full-term development after ICSI.     

Survival of bovine embryos stored at 4 degrees C

These experiments were designed to test the efficacy of storing bovine embryos at 4 degrees C. Of particular interest were the age of embryo at which maximum post-storage survival could be achieved and longevity at 4 degrees C. A greater proportion of day 8 blastocysts developed in vitro at 37 degrees C following refrigeration for 48 hr than did embryos collected 2, 4 or 6 days after estrus (P<0.01). Survival of blastocysts stored at 4 degrees C for 48 hr was similar to that of nonstored blastocysts. In a subsequent experiment, day 8 blastocysts were recovered nonsurgically and assigned to one of the following treatments: (a) immediate transfer; (b) culture at 37 degrees C; or (c) storage at 4 degrees C for 1, 2, 3 or 5 days. Post-storage viability was assessed by either development in culture at 37 degrees C or embryo survival following nonsurgical transfer to synchronized recipients. In vitro survival of nonstored embryos and embryos stored 1 day did not differ. Survival decreased after storage for 2 days (P<0.10) or longer (P<0.05). Similar results were observed for survival after transfer, but embryo viability decreased even more rapidly with increasing duration of storage. In vitro survival was approximately 50% for blastocysts stored for 3 and 5 days, but few pregnancies resulted from transfer of embryos stored for these periods. In another experiment survival after transfer of blastocysts stored at 4 degrees C for up to 2 days was similar to that of nonstored blastocysts.     

The microbial association of Greek taverna sausage stored at 4 and 10 degrees C in air, vacuum or 100% carbon dioxide, and its spoilage potential

Strains of the Lactobacillus sakei/curvatus group, mainly non-slime-producing Lact. sakei, dominated the microbial flora of industrially manufactured taverna sausage, a traditional Greek cooked meat, stored at 4 degrees C and 10 degrees C in air, vacuum and 100% CO2. Atypical, arginine-positive and melibiose-negative strains of this group were isolated. The isolation frequency of Lact. sakei/curvatus from sausages stored anaerobically was as high as 92-96%, while other meat spoilage organisms were practically absent. Conversely, in air-stored sausages, leuconostocs, mainly Leuconostoc mesenteroides ssp. mesenteroides, had a considerable presence (14-21%), whereas Brochothrix thermosphacta, pseudomonads and Micrococcaceae grew, but failed to increase above 10(5) cfu g(-1) in all samples during storage. Only yeasts were able to compete against LAB and reached almost 10(7) cfu g(-1) after 30 d of aerobic storage at 10 degrees C. The great dominance (> 10(8) cfu g(-1)) of LAB caused a progressive decrease of pH and an increase of the concentration of L-lactate, D-lactate and acetate in all sausage packs. The growth of LAB and its associated chemical changes were more pronounced at 10 degrees C than 4 degrees C. At both storage temperatures, L-lactate and acetate increased more rapidly and to a higher concentration aerobically, unlike D-lactate, which formed in higher amounts anaerobically. Storage in air was the worst packaging method, resulting in greening and unpleasant off-odours associated with the high acetate content of the sausages. Carbon dioxide had no significant effect on extending shelf-life. The factors affecting the natural selection of Lact. sakei/curvatus in taverna sausage are discussed. Moreover, it was attempted to correlate the metabolic activity of this group with the physicochemical changes and the spoilage phenomena occurring in taverna sausage under the different storage conditions.     

The effect of pH on the viability of hypothermically stored rat hepatocytes

Hepatocytes isolated from the rat liver were stored for up to 72 hr at 4 degrees C in a tissue culture medium (Liebovitz-15) at different pH values to determine how pH affects hepatocyte viability. This is a model to simulate cold storage of livers for transplantation and determine the optimal pH for maintenance of liver cell function. The cells were stored in the absence of oxygen. At the end of cold storage the percentage of the total cellular LDH released into the extracellular medium was used as a measure of hepatocyte viability. Also, lactate dehydrogenase (LDH) release was determined in hepatocytes incubated at normothermia (37 degrees C) for 90 min following 72 hr of cold storage. The results demonstrate that hepatocytes tolerate a wide range of pH values in the storage medium and that only about 10% of the total LDH was released from hepatocytes stored up to 72 hr at pH's from 5.0 to 8.0. Normothermic incubation, however, demonstrated that the pH of the storage medium affected viability. After 48 hr of storage only hepatocytes stored at pH values from 7.0 to 8.0 remained viable (LDH release similar to that of freshly incubated hepatocytes = 28 +/- 7.2%). After 72 hr of storage and 90 min of normothermic incubation, hepatocytes incubated at all pH values studied were nonviable (greater than 60% release of LDH). These results suggest that the optimal pH for storage of hepatocytes at 4 degrees C is near neutrality (7.0 to 7.4).     

Viability of nuclei of two-cell mouse embryos stored at 4 degrees C and fused with blastomeres of fresh two-cell embryos.

This study compares the resistance of the nuclei and the cytoplasm of two-cell mouse embryos to short-term storage at low temperature above 0 degrees C. Two-cell embryos were stored at 4 degrees C for 24-96 h in PB1 containing 0.25, 0.5, 0.75, and 1.0 M sucrose. The development to blastocysts in culture was highest in the presence of 0.5 M sucrose. However, only 3% of the embryos developed into blastocysts after 96 h of storage. On the other hand, the viability of the nuclei of two-cell embryos stored at 4 degrees C was significantly prolonged when they were transplanted into a blastomere of enucleated fresh F1 (C57BL/6JXCBA) two-cell embryos. The proportions of chimeric embryos that developed to blastocysts were 88, 67, 76, 71, 64, 45, 32, and 20% following storage for 0, 48, 72, 96, 120, 144, 168, and 192 h, respectively. In addition, there was no difference in the coat color of the young derived from nuclei stored at 4 degrees C or fresh nuclei, although the proportions of chimeric embryos that developed into live young after transfer tended to decrease with increased storage time. Moreover, the viability of nuclei stored at 4 degrees C for 192 h was confirmed in the germ cell population of chimeric mice mated with albino mice. These results demonstrated that the nuclei in the two-cell mouse embryos were more resistant to storage at low temperature than the cytoplasm.     

Inhibition of colloid cell swelling in rabbit kidney cortex by disodium glycerophosphate.

Solution based on disodium glycerophosphate have been assessed for ability to inhibit cellular swelling during hypothermic storage of slices of rabbit renal cortex. It was found that slices stored in glycerophosphate media for 3 days gained about 50% of their control weight, whereas control slices stored in chloride based media gained 50% of their control weight after only 2 hr of storage and gained about 128% of their initial weight after 3 days. Electrolyte content of slices stored at 4 °C reflected the change in hydration. When slices were warmed after various storage times, it was found that viability was much better preserved in glycerophosphate saline than in chloride based saline, although the slices stored in glycerophosphate media contained more sodium prior to warming than slices stored in conventional media. Glycerophosphate solutions appear to be suitable for routine use in experiments with renal cortical slices in the vicinity of 0 °C when cell volumes are to be held constant for prolonged periods.     

Development, DNA fragmentation and cell death in porcine embryos after 24 h storage under different conditions

For practical applications of porcine embryo transfer (ET) it is important to develop feasible embryo storage conditions. The aim of the present study was to evaluate the effect of short-term storage (24 h) on the quality of in vivo produced porcine embryos. Three temperatures 18, 25 and 38 degrees C and three different media: Dulbecco's phosphate buffered saline (DPBS), TCM199 and Emcare, were tested for two different embryo ages: D4 embryos (collected 144 h after hCG treatment) and D5 embryos (collected 168 h after hCG). After slaughter of the donor gilts, embryos were collected and transported at 25 degrees C to the lab where morulas and blastocyst were selected (D4 n = 222; D5 n = 167) and randomly used as controls or distributed over the treatment groups. Developmental stage and embryo diameter were assessed by normal light microscopy, while total number of cells and incidence of apoptosis were assessed using a fluorescent embryo quality staining technique that combines three different dyes: Ethidium Homodimer (EthD-1), TUNEL and Hoechst 33342. Following 24 h storage, D5 embryos had higher rates of hatching (24%) and degeneration (9%) compared to D4 embryos (10 and 4%, respectively; P < 0.05). Embryos stored at 38 degrees C had higher rates of hatching (37%) compared to those ones stored at 25 degrees C (13%) or 18 degrees C (0%; P < 0.01). More embryos hatched when stored in medium Dulbecco's phosphate buffered saline (DPBS) or in TCM199 compared to those stored in Emcare (P < 0.05). A higher percentage of embryos stored at 18 degrees C degenerated compared to those stored at 25 or 38 degrees C (P < 0.01). No significant increase in apoptosis was observed after storage compared to the rates of apoptosis at 0 h (controls) or between the different storage groups. Based on the results we conclude that D4 porcine embryos produced in vivo, selected under normal light microscopy and stored at 25 degrees C in a serum free medium for 24 h will have a suitable developmental stage for ET and a high embryo quality.     

Viability and Endogenous Substrates Used During Starvation Survival of Rhodospirillum rubrum

Cells of Rhodospirillum rubrum were grown photoorganotrophically and chemoorganotrophically and then starved for organic carbon and combined nitrogen under four conditions: anaerobically in the light and dark and aerobically in the light and dark. Illumination prolonged viability and suppressed the net degradation of cell material of phototrophically grown cells, but had no effect on chemotrophically grown cells that did not contain bacteriochlorophyll. The half-life survival times of carbohydrate-rich phototrophically grown cells during starvation anaerobically or aerobically in the light were 17 and 14.5 days, respectively. The values for starvation aerobically and anaerobically in the dark were 3 and 0.5 days, respectively. Chemotrophically grown cells had half-life survival times of 3 and 4 days during starvation aerobically in the light and dark, respectively, and 0.8 day during starvation anaerobically in the light or dark. Of all cell constituents examined, carbohydrate was most extensively degraded during starvation, although the rate of degradation was slowest for phototrophically grown cells starved anaerobically in the light. Phototrophically grown cells containing poly-beta-hydroxybutyrate as carbon reserve were less able to survive starvation anaerobically in the light than were carbohydrate-rich cells starved under comparable conditions. Light intensity had a significant effect on viability of phototrophically grown cells starving anaerobically. At light intensities of 320 to 650 lx, the half-life survival times were 17 to 24 days. At 2,950 to 10,500 lx, the survival times decreased to 1.5 to 5.5 days. The kinetics of cell death correlated well with the rate of loss of cell mass of starving cells. However, the cause of death could not be attributed to degradation of any specific cell component.     

Studies on the endogenous metabolism and senescence of starved Sarcina lutea

1. When washed suspensions of Sarcina lutea are starved aerobically in phosphate buffer at the growth temperature of 37 degrees , the rate of endogenous oxygen consumption decreases to very low values after 10hr., although many of the cells survive for 40hr. If starvation is prolonged further, the bacteria die at a rate of approximately 1.5% of the initial viable population per hour. 2. Oxidation of intracellular free amino acids accounts for most of the observed endogenous oxygen uptake but RNA is also utilized and a portion of the component bases and pentose is degraded and presumably oxidized. Ammonia appears in the supernatant and some pentose and ultraviolet-absorbing nucleotide are released from the cells. DNA, protein and polysaccharide are not measurably degraded. 3. Survival can be correlated with the ability of aerobically starved bacteria to oxidize exogenous l-glutamate and glucose. When starved under nitrogen for 40hr. cells continue to oxidize their endogenous reserves at undiminished rates when transferred to aerobic conditions; on prolonging anaerobic starvation the rate of oxidation declines during the period of most rapid loss of viability. 4. In the presence of Mg(2+), RNA degradation during aerobic starvation is almost completely suppressed without affecting the period for which the bacteria survive. 5. Cells grown in peptone supplemented with glucose accumulate reserves of polysaccharide which are metabolized in aerobic starvation, together with free amino acids. Ammonia is evolved and RNA is degraded to a greater extent than in peptone-grown suspensions. Bacteria rich in polysaccharide survive less well than those which are deficient in the polymer; the reason for this phenomenon has yet to be established. 6. In peptone medium, endogenous oxygen uptake and the concentration of intracellular free amino acids decline as growth progresses and they continue to decrease when the organism is held in stationary phase. Under the conditions used, the endogenous Q(o2) and free amino acid pool of cells grown in peptone with 2% (w/v) glucose did not decline so markedly and the bacteria contained large amounts of polysaccharide at all stages of growth.     

Effect of anaerobiosis on the adult cerambycid beetle, Rhagium inquisitor L.

Glycerol, lactate, and alanine were found accumulated in adult Rhagium inquisitor beetles which had been incubated anaerobically at +5°C for one month. However, glycerol and alanine were accumulated also in control beetles, kept aerobically at the same temperature, indicating that the accumulation of these substances is an effect of the low temperature. Lactate was rapidly destroyed when the beetles were transferred to air, and this process was accompanied by increased oxygen consumption, indicating the repaying of an oxygen debt due to lactate accumulation. This high oxygen consumption phase was followed by a second, more prolonged high oxygen consumption phase. Because this pattern of oxygen consumption was not found in the control beetles, the second phase of high oxygen consumption was also probably due to the anoxia. It is suggested that this increased oxygen consumption might be caused by a metabolic destruction of accumulated metabolites, but no substances have been identified which could account for it. For this reason the elevated oxygen consumption is more likely to be due to an extensive protein synthesis, related to the re-establishing of aerobic enzyme systems after the long-lasting anoxia.     

Determinants in microbial colonization of the murine gastrointestinal tract: pH, temperature, and energy-yielding metabolism of Torulopsis pintolopesii.

Torulopsis pintolopesii is an indigenous yeast that colonizes the secreting epithelia in the stomachs of mice and rats. A wild-type strain of this microbe was isolated and identified. To attempt to learn characteristics of the yeast that are advantageous to it in colonizing its natural habitat in vivo, we examined some aspects of its nutrition and energy-yielding metabolism and some environmental conditions that influence its growth in vitro. The yeast appeared to be limited in the compounds it can utilize as carbon and nitrogen sources. It grew best at 37 degrees C and did not grow at 23 or 43 degrees C. It grew optimally at neutral pH but could grow aerobically at pH values as low as 2.0 and anaerobically at pH values as low as 3.4. As assessed by measurements of growth rates and yield coefficients, it grew better aerobically than anaerobically. When grown aerobically, it had a cyanide-sensitive system for taking up O(2) and tested positively for cytochrome c oxidase activity. A petite mutant strain isolated from the wild-type strain had a growth rate and yield coefficient when incubated aerobically that were essentially the same as those of the wild-type parent grown anaerobically. Likewise similar to the wild-type parent grown anaerobically, the petite strain, though incubated aerobically, did not take up O(2). Yeast-free mice associated with either the wild-type or the petite mutant strain were colonized at essentially the same rates and to similar final population levels by both strains. The yeast's capacity to respire may be of little advantage to it in its natural environment. By contrast, its abilities to grow best at 37 degrees C and to grow at low pH values are undoubtedly advantageous characteristics in this respect. The limitations in its carbon and nitrogen nutrition are difficult to evaluate as ecological factors in its colonization of the natural habitat.