Biological activity of Friend spleen focus-forming virus after cold storage

Two stocks of Friend spleen focus-forming virus (SFFV) were prepared, one in saline and the other in Eagle's medium with 2% fetal calf serum, and the effects of different freezing, storage and thawing temperatures were determined for the recovery of infectious virus from each diluent. Once frozen, virus maintained its titer at ?70 and at ?170 °C for up to 13 weeks, while it lost titer at ?13 °C more rapidly if it had been prepared in saline than in medium. However, during the freezing process lower ambient temperatures (?70 and ?170 °C) gave lower virus yields than a higher temperature (?13 °C) did. Similarly, rapid thawing (in a 37 °C water bath) was less efficient than slow thawing (in 4 or 20 °C air) for the recovery of infectious SFFV, This study illustrates the importance, for efficient recovery of leukemogenic activity from stored murine leukemia virus stocks, of the temperature used for freezing or thawing, as well as for storage.     

Strong light inhibits germination of Artemisia sphaerocephala and A. ordosica at low temperature and its relevance to revegetation in sandy lands of Inner Mongolia, China

Artemisia sphaerocephala and A. ordosica are two dominant shrub species in Mu Us sandy land (Inner Mongolia, China) and are widely used for vegetation restoration. However, there are two different conclusions about the effect of light on their germination: light promotes germination versus light inhibits germination. The aim of this study was to evaluate these two conclusions and relate the results to instances of failure of these two species to germinate well when air-dispersed in revegetation projects. The effects of fluctuating temperature, light/dark, source (population), position on mother plant, storage condition, and storage time were tested on germination of achenes of these two species. At low temperature, final percent germination (FPG) of achenes in dark and nearly dark conditions was significantly higher than those in light. At 10:20°C, achenes of both A. sphaerocephala and A. ordosica had higher FPG in dark than in light regardless of source, position on mother plants or storage condition. At suboptimum (5:15°C) and supraoptimal (25:35°C) temperatures, germination of A. sphaerocephala and A. ordosica achenes was inhibited in both light and darkness. It was concluded that light inhibits germination of A. sphaerocephala and A. ordosica achenes at low (10:20°C) temperature but not at high (15:25°C) temperature. Since the temperature in Mu Us sandy land is around 10:20°C in early June, when air sowing is done, achenes should germinate best when they are covered by a thin layer of sand.     

Preservation of energy-linked functions of insect mitochondria by simple freezing methods.

Attempts are made to preserve some energy-linked functions of mitochondria isolated from the fat body of the cockroach Nauphoeta cinerea. The ability of mitochondria to sustain appreciable levels of oxidation, phosphorylation, respiratory control and 2, 4-DNP-stimulation of respiration which are normally lost during incubation at 30°C for 2 hrs or during aging at 0–2°C for 48 hrs can be preserved effectively by storing mitochondrial suspensions in sucrose - EDTA medium for 1 week at - 20°C and at least 3 weeks at -196°C (liquid nitrogen temperature). DMSO (dimethyl sulfoxide; 10% v/v) was found to be ineffective as an aid to preservation at 0–2°C, a significant aid at -20°C and unnecessary at -196°C. With slight variations (for flight muscle mitochondria which requires DMSO at -196°C), this procedure is also effective for mitochondria isolated from other tissues of various insect species. EDTA was found to be an essential ingredient of the isolation medium and therefore for all storage procedures. Both lyophilization and the preincubation of mitochondria with nupercaine (400 μM) have deleterious effects.     

Collagenolytic activity in several species of deuteromycetes under various storage conditions

The ability of deuteromycetes of the genera Penicillium, Aspergillus, and Botrytis to retain collagenolytic activity was studied after both 2 and 10 years of storage on a Czapek medium under a layer of mineral oil at 4°C, as well as in silica gel granules at 20 and ?60°C. The enzymatic activity of several species, including Botrytis terrestris, Penicillium janthinellum, Penicillium chrysogenum, and Penicillium citrinum, was retained under both conditions of storage. Aspergillus repens retained enzymatic activity only if stored under a layer of mineral oil. The viability of conidia and the collagenolytic activity of Botrytis terrestris, P. janthinellum, P. chrysogenum, and Penicillium citrinum, maintained on silica gel for 10 years, depended on the storage temperature. The viability of the test strains improved after storage on a silica gel at ?60°C. A strain of Aspergillus repens lost its ability to dissolve collagen at various storage tempeatures on the silica gel. The index of lysis for three strains of Penicillium deuteromycetes (Penicillium janthinellum, Penicillium chrysogenum, and Penicillium citrinum) increased after a 10-year storage on silica gel at ?60°C.     

Mortality of bifidobacteria in boiled yogurt

Bifidobacterial strains showed various mortalities during storage at 30°C in boiled yogurt prepared with Streptococcus sp. and Lactobacillus sp. Bifidobacterium breve 203 was most stable, maintaining its initial cell number for more than 5 days. It was also stable at 4 or 10°C in fresh yogurt. B. longum 401 rapidly lost viability in the boiled yogurt and was unstable in the fresh yogurt at any temperature. Lactobacillus sp. remained fully viable for one week or more at 4–30°C while Streptococcus sp. lost viability at 20°C or above. The difference in mortality between B. breve 203 and B. longum 401 was mainly due to their sensitivity to the acidic environment, with temperature during storage having a secondary effect. Effects of lysozyme, pepsin and bile acids on the two strains were also investigated.     

Survival,haemolymph osmolality and tissue water of Penaeus chinensis juveniles acclimated to different salinity and temperature levels

Survival, growth, haemolymph osmolality and tissue water of Penaeus chinensis (Osbeck) juveniles (0.11 ± 0.04 g) were investigated, after they were acclimated to 10, 20, 30 and 40 ppt from 33 ppt for 14 days at 24°C, and then acclimated to 12, 18, 24 and 30°C at each salinity for 14 days. The survival of shrimp was the lowest at 10 ppt and 12°C. Growth of shrimp increased with increased temperature in the range 12–24°C, with no significant difference among four salinity levels at 18, 24 and 30°C. Haemolymph osmolality increased with increased salinity, and decreased with increased temperature. The isosmotic point computed from the linear relationship between haemolymph osmolality and medium osmolality was 664, 632, 629 and 602 mOsm/kg which is equivalent to 25.2, 24.1, 24.0 and 23.1 ppt at 12, 18, 24 and 30°C, respectively. Tissue water decreased with increased medium osmolality and haemolymph osmolality. The slope obtained from the relationship between haemolymph osmolality and medium osmolality indicated that there is an impairment of osmoregulatory ability for the P. chinensis juveniles at 12°C.     

n-3 Fatty acid content in eggs laid by hens fed with marine algae and sardine oil and stored at different times and temperatures

Inclusion of sardine oil (SO) in diets for laying hens significantly increases the n-3 polyunsaturated fatty acids (PUFAs) in the egg, but these are more sensitive to oxidation, so the storage time and temperature can cause a decrease in their concentration. Therefore, the objective of this study was to determine the effect of algae Macrocystis pyrifera, Enteromorpha spp., and Sargassum sinicola on n-3 PUFA contents in eggs from laying hens fed diets supplemented with sardine oil and stored for different times (0, 15, and 30 days) and temperatures (20°C and 4°C), for 8 weeks. One hundred and twenty hens were divided into four treatments: T1 (commercial diet), T2 (2% SO?+?10% M. pyrifera), T3 (2% SO?+?10% Enteromorpha), and T4 (2% SO?+?10% S. sinicola). At the end, 50 eggs per treatment were collected to quantify total lipids and egg n-3 PUFAs at different times (0, 15, and 30 days) and temperatures (20°C and 4°C) of storage. The results were analyzed using a 3?×?3?×?2 factorial design, and Tukey test to compare means (P?<?0.05). The results show that M. pyrifera and S. sinicola had a better effect on eicosapentaenoic acid, while Enteromorpha was better for docosahexaenoic acid. In relation to time and temperature, the content of the fractions analyzed in the three treatments at 15 days/4°C had a lower loss compared with eggs analyzed at day 0/20°C.     

Produktion von Ochratoxin A und B durch<Emphasis Type="Italic">Penicillium verrucosum</Emphasis> auf Gerste in Abhängigkeit der Parameter Wasseraktivität,Wassergehalt und Temperatur

The ochratoxin A and B (OTA, OTB) production by a toxigenic isolate ofPenicillium verrucosum grown on brewing barley up to six weeks was studied at a storage temperature of 25 °C and different moisture and water activity conditions. Sorption isothermes for barley were prepared at temperatures of 10°C, 15°C and 25°C. OTA was produced after 2 weeks of storage at moisture contents of ≥19%, which is equivalent to water activities (a_w) of 0.83 (adsorptive) and 0.82 (desorptive) at 25 °C. Increased OTA concentrations (5.8-fold and 16.1-fold) were noticed when the moisture contents were adjusted to 20% (a_{w [ads]} 25 °C=0.86) and 21% (a_{w [ads]} [ 25 °C=0.88), respectively. An increase was also shown during storage of 4 and 6 weeks (1.2-fold and 2.4-fold, respectively). Production of OTB was shown to occur at moisture contents ≥18% (a_{w [ads]} 25 °C=0.81). The findings document that OTA and OTB are not produced byP. verrucosum grown on barley stored below 18% moisture content.     

The influence of temperature, relative humidity and host factors on the attachment and survival of Boophilus microplus (Canestrini) larvae to skin slices

Attachment of B. microplus larvae was examined using slices (0.5 mm) of bovine skin stretched over a suitable medium. Optimal temperature for attachment lay between 31 and 38°C. At 38°C, 70 to 80% had attached by 4 h but this was followed 6–7 h after the release of the larvae by a significant decrease in the percentage attached. By 8 h, the percentage attached had risen again and remained at 70–80% for the subsequent 16 h. Attachment was not influenced by ambient relative humidity within the range 20–75 % on the time scale studied (16 h). Most larvae denied access to the skin surface at 20 or 45 % R.H. (38°C) died within 24 h. Those allowed to feed survived at 45% but most died at 20% R.H. There was no difference in attachment when bovine or rabbit serum or phosphate buffered saline was used, nor when skin was taken from Zebu × European cattle or Herefords. Attachment was no different when skin was taken from neck, rib or rump, or from cattle with different levels of resistance to B. microplus but there was reduced attachment with mouse skin. Attachment was reduced on skin which had been stored at ? 15°C for 1 day but storage at 10°C for 16 h had no effect. These results are related to problems of tick water balance, host specificity and stimuli for tick attachment.     

Effect of Environmental Temperatures on Proteome Composition of Salmonella enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium (STM) is a major cause of gastroenteritis and transmitted by consumption of contaminated food. STM is associated to food originating from animals (pork, chicken, eggs) or plants (vegetables, fruits, nuts, and herbs). Infection of warm-blooded mammalian hosts by STM and the underlying complex regulatory network of virulence gene expression depend on various environmental conditions encountered in hosts. However, less is known about the proteome and possible regulatory networks for gene expression of STM outside the preferred host. Nutritional limitations and changes in temperature are the most obvious stresses outside the native host. Thus, we analyzed the proteome profile of STM grown in rich medium (LB medium) or minimal medium (PCN medium) at temperatures ranging from 8 °C to 37 °C. LB medium mimics the nutritional rich environment inside the host, whereas minimal PCN medium represents nutritional limitations outside the host, found during growth of fresh produce (field conditions). Further, the range of temperatures analyzed reflects conditions within natural hosts (37 °C), room temperature (20 °C), during growth under agricultural conditions (16 °C and 12 °C), and during food storage (8 °C). Implications of altered nutrient availability and growth temperature on STM proteomes were analyzed by HPLC/MS-MS and label-free quantification. Our study provides first insights into the complex adaptation of STM to various environmental temperatures, which allows STM not only to infect mammalian hosts but also to enter new infection routes that have been poorly studied so far. With the present dataset, global virulence factors, their impact on infection routes, and potential anti-infective strategies can now be investigated in detail. Especially, we were able to demonstrate functional flagella at 12 °C growth temperature for STM with an altered motility behavior.     

Impact of storage conditions and duration on function of native and cargo-loaded mesenchymal stromal cell extracellular vesicles

Background aimsAs evidenced by ongoing clinical trials and increased activity in the commercial sector, extracellular vesicle (EV)-based therapies have begun the transition from bench to bedside. As this progression continues, one critical aspect of EV clinical translation is understanding the effects of storage and transport conditions. Several studies have assessed the impact of storage on EV characteristics such as morphology, uptake and component content, but effects of storage duration and temperature on EV functional bioactivity and, especially, loaded cargo are rarely reported.MethodsThe authors assessed EV outcomes following storage at different temperatures (room temperature, 4°C, –20°C, –80°C) for various durations as well as after lyophilization.ResultsMesenchymal stromal cell (MSC) EVs were observed to retain key aspects of their bioactivity (pro-vascularization, anti-inflammation) for up to 4–6 weeks at –20°C and –80°C and after lyophilization. Furthermore, via in vitro assays and an in vivo wound healing model, these same storage conditions were also demonstrated to enable preservation of the functionality of loaded microRNA and long non-coding RNA cargo in MSC EVs.ConclusionsThese findings extend the current understanding of how EV therapeutic potential is impacted by storage conditions and may inform best practices for handling and storing MSC EVs for both basic research and translational purposes.     

Suitability of poor medium in counting total viable airborne bacteria

The purpose of this study was to test the effect of incubation temperature and culture medium on viable counts of airborne bacteria. The incubation temperature had different effect on indoor and outdoor air bacteria. Indoor air bacteria grew as well at 20°C as 37°C, but less at 10°C. Outdoor air bacteria grew equally well at 10°C and 20°C, but less at 37°C. Both indoor and outdoor air bacteria grew differently on poor and rich media. The counts of both indoor and outdoor air bacteria were higher on poor R2A medium (low nutrient concentration) than on rich TYG and blood media (high nutrient concentration). The results indicate that a poor medium incubated at 20°C is adequate for counting viable airborne bacteria.     

In vitro conservation of enset under slow-growth conditions

Studies on in vitro storage of enset under slow-growth conditions were carried out to develop an efficient protocol for conservation of the genetic diversity of the crop. The response to different growth retardation treatments was examined using three enset clones collected from southwestern Ethiopia. In vitro cultures could be effectively maintained for 6 months at 15 °C and 18 °C on MS medium supplemented with 10 μM BAP, in the presence of mannitol at concentrations of 0, 1 or 2% as a growth retardant. Shoots were subsequently recovered and multiplied on MS medium supplemented with 10 and 20 μM BAP at 25 °C and rooted shoots were successfully transferred to the greenhouse. Incubation at the lower temperature (15 °C) and the presence of mannitol in the culture medium had a significantly positive effect on maintenance, measured by the number of recovered shoots after storage. This revised version was published online in June 2006 with corrections to the Cover Date.     

Stem elongation of ornamental bromeliad in tissue culture depends on the temperature even in the presence of gibberellic acid

Acanthostachys strobilacea Link, Klotzsch, & Otto is an ornamental bromeliad native to Brazilian Atlantic Forest that naturally exhibits a rosette growth pattern. According to the temperature conditions of the in vitro culture, this species can exhibit stem elongation, facilitating the isolation of the nodal segments to be applied in its micropropagation. The rosette morphology is reestablished when this species is maintained under low temperature, thus allowing the maintenance of a germplasm collection (slow growth storage). Gibberellins (GA) are usually applied to stimulate stem elongation in micropropagated plants. Thus, our aim here was to verify the influence of temperature over the stem elongation of A. strobilacea when GA₃ is applied to the medium, thus estimating the use of this phytoregulator in slow growth cultures at low temperatures. Physiological and anatomical studies were performed on plants obtained from nodal segments maintained at 10, 15, 20, and 25 °C. Regardless of the applied treatment, no segments developed at 10 °C. Stem elongation occurred at 25 and 30 °C, and was not seen for plants grown under 15 and 20 °C. The application of 50 µM of GA₃ restored stem elongation in plants at 20 but not at 15 °C. The influence of gibberellins on stem elongation of this tropical bromeliad depends on the cultivation temperature, in which low temperature preponderates over the stem elongation effects of GA₃. In addition, the optimum temperature for the slow growth of this species depends on the starting temperature of the explant used in the micropropagation.     

Germination responses of four native terrestrial orchids from south-west Western Australia to temperature and light treatments

We report an investigation into the impact of temperature and illumination on in vitro symbiotic and asymbiotic germination of the threatened taxon Caladenia huegelii, and three other orchid spp. namely—Caladenia latifolia, Microtis media and Pterostylis sanguinea, all species from south-west Western Australia, a recognized biodiversity hotspot. High symbiotic germination on oatmeal agar (OMA + fungal symbionts specific to each species) was recorded in three species in continuous dark incubation i.e. C. huegelii seeds (98 % germination at 25 °C), and M. media and P. sanguinea (93 and 98 % respectively at 20 °C). Highest symbiotic germination for C. latifolia (100 %) was observed at 15 and 20 °C under light treatment (12/12 h light/dark). Low temperature incubation (10 °C) significantly suppressed symbiotic germination/development of seedlings across all species. Asymbiotic media treatments assessed (OMA minus fungal symbionts, Pa₅ and ½ MS), failed to stimulate any germination with C. latifolia seeds at 20 °C in either light or dark treatments after an 8 week incubation period. Seeds of M. media sown onto ½ MS medium resulted in higher germination in all developmental stages (3–5) in dark treatment than OMA and Pa₅. Seeds of P. sanguinea sown onto ½ MS medium resulted in higher overall germination in all developmental stages (3–5) in light and dark incubation compared to OMA and Pa₅. OMA supported the highest asymbiotic germination (100 %) in both light and dark incubation with M. media (only to stage 3) but did not support germination and development with other spp. tested. Caladenia huegelii seeds reached developmental Stage 3 (i.e. germinated), but only on Pa₅ medium and only at a relatively low rate in either light (2.6 %) or dark (2.1 %). Germination was higher and development of seedlings faster overall in all test species in symbiotic compared with asymbiotic media treatments. P. sanguinea seeds demonstrated the best response (among species tested) to asymbiotic germination on ½ MS with 40–53 % of germinated seeds spread over developmental stages 3–5 in light or dark incubation (at 20 °C) respectively. Illumination had no effect on fungal symbiont growth across all species, however incubation temperature treatments (10, 15, 20 and 25 °C) affected fungal growth rate. Growth of the fungal symbionts of C. huegelii, M. media and C. latifolia demonstrated significantly lower activity at 10 °C, but the cumulative radial growth rate of the P. sanguinea fungal symbiont reached 64 cm² after only 2 weeks at all temperatures tested, including 10 °C. The study highlights differences in symbiotic and aysmbiotic germination and early protocorm development in vitro between co-occurring herbaceous terrestrial Australian orchid taxa in response to variations in basal media, temperature and light.     

Low temperature alters plasma membrane lipid composition and ATPase activity of pineapple fruit during blackheart development

Plasma membrane (PM) plays central role in triggering primary responses to chilling injury and sustaining cellular homeostasis. Characterising response of membrane lipids to low temperature can provide important information for identifying early causal factors contributing to chilling injury. To this end, PM lipid composition and ATPase activity were assessed in pineapple fruit (Ananas comosus) in relation to the effect of low temperature on the development of blackheart, a form of chilling injury. Chilling temperature at 10 °C induced blackheart development in concurrence with increase in electrolyte leakage. PM ATPase activity was decreased after 1 week at low temperature, followed by a further decrease after 2 weeks. The enzyme activity was not changed during 25 °C storage. Loss of total PM phospholipids was found during postharvest senescence, but more reduction was shown from storage at 10 °C. Phosphatidylcholine and phosphatidylethanolamine were the predominant PM phospholipid species. Low temperature increased the level of phosphatidic acid but decreased the level of phosphatidylinositol. Both phospholipid species were not changed during storage at 25 °C. Postharvest storage at both temperatures decreased the levels of C18:3 and C16:1, and increased level of C18:1. Low temperature decreased the level of C18:2 and increased the level of C14:0. Exogenous application of phosphatidic acid was found to inhibit the PM ATPase activity of pineapple fruit in vitro. Modification of membrane lipid composition and its effect on the functional property of plasma membrane at low temperature were discussed in correlation with their roles in blackheart development of pineapple fruit.     

Predicting the stabilities of freeze-dried suspensions of Lactobacillus acidophilus by the accelerated storage test

The elevated temperatures of 50 ° C, 60 ° C, and 70 ° C were used in an accelerated storage test for predicting the stability of freeze-dried suspensions of L. acidophilus. The logarithmic death of bacteria at the above temperatures and the Arrhenius relationship obtained permitted predicting the rate of death at any storage temperature. The values predicted for storage stability of freeze-dried suspensions of L. acidophilus at 4 ° C and 20 ° C were confirmed by the actual values obtained after storage at these temperatures for 6, 15, and 19 mo.     

Changes in growth and size of Keratella cochlearis (Gosse) in relation to some environmental factors in cultures

Keratella cochlearis (Gosse) was cultured non-axenically in Carefoot medium diluted with Erken water at 5 °C, 15 °C and 20 °C with Rhodomonas minuta (Skuja) as a food alga. The rotifer reached ca. 120 ind. ml^?1, having generation times of 2–7 days, a Q₁₀-value of ca. 2, and at the lowest temperature >20% longer posterior spines. When co-cultured with Chlorella sp., at 0–30 mg Ca l^?1 and 1.6 meq NaHCO₃ l^?1 in medium L 11 at 20 °C, the maximum generation time and individual numbers were 3–4 days and up to 100 ind. ml^?1, respectively. Animal numbers increased in relation to nutrient multiples, up to two multiples, of the culture medium L 16. Growth and length were reduced, although the width increased above two multiples of this culture medium. The trace metal tolerance was broad and increased additions of a metal mixture (L 11) slightly increased the length of the rotifers. No major changes in the length were observed when HCO₃ or Ca were varied in the culture medium (L 11), although a decrease in the length was noted in old cultures.     

In vitro conservation of chestnut (Castanea sativa) by slow growth

Slow growth storage has been achieved for Castanea sativa (cv. ‘Montemarano’) shoot cultures over a duration of 48 mo at a temperature of 8°C, where 82% of explants survived and were able to resume normal growth after transfer to standard culture conditions at 23°C. The evaluation of the chlorophyll content of leaves also showed no differences between material stored for 48 mo and control material subcultured at 23°C. With a storage temperature of 4°C, the survival of shoots was significantly lower at approximately 56% after 12 mo, and no plants recovered after 24-mo storage. The presence of 6-benzyladenine 0.44 μM in the culture medium proved to be necessary for the recovery of healthy shoots, while pre-treatments with different concentrations of abscisic acid did not significantly influence the survival of shoots following storage conditions. A low level of light during slow growth storage resulted in positive effects on the rate of shoot survival over the longest preservation periods.     

Neutrophil migration in canines: In vivo comparison of granulocyte function following 24-hour storage at 6 or 20 °C of leukocyte concentrates or of granulocytes purified by counterflow centrifugation-elutriation

The use of granulocyte-rich concentrates from leukapheresis purified by counterflow centrifugation—elutriation to obtain pure granulocytes for transfusion studies in cyclo-phosphamide-induced neutropenic animal models is reported. Our data for granulocyterich leukapheresis concentrates indicate that room temperature (20 °C) appears to be preferred to 6 °C for short-term granulocyte storage. The data also indicate that although the granulocytes isolated by counterflow centrifugation—elutration may retain in vitro functions of chemotaxis, phagocytosis, and bactericidal activity, the in vivo function of migration into skin chambers for isolated granulocytes is seriously impaired after storage for 18 to 24 hr at both 6 and 20 °C. This loss of in vivo function of stored granulocytes occurs in isolated granulocytes obtained by both counterflow centrifugation-elutriation and dextran sedimentation, and it is not observed in the leukocyte concentrates held at 20 °C. The results of these studies are fourfold. First, freshly isolated granulocytes display no apparent loss of either in vivo or in vitro function. Second, granulocytes isolated by counterflow centrifugation-elutriation or dextran sedimentation and stored at 6 or 20 °C are severely impaired in terms of their in vivo chemotactic function but display no loss of in vitro efficacy. Third, 20 °C storage of granulocyte-rich leukapheresis concentrates for 18 to 24 hr is superior to 6 °C storage. Fourth, in vitro analysis may be limited in its ability to indicate in vivo function as a measure of success in granulocyte preservation studies.