Effect of incubation temperature on isolation of Campylobacter jejuni genotypes from foodstuffs enriched in Preston broth

Preston broth and agar incubated at either 37 or 42 degrees C have been widely used to isolate campylobacters from foodstuffs. The consequences of using either incubation temperature were investigated. Retail packs of raw chicken (n = 24) and raw lamb liver (n = 30) were purchased. Samples were incubated in Preston broth at 37 and 42 degrees C and then streaked onto Preston agar and incubated as before. Two Campylobacter isolates per treatment were characterized. Poultry isolates were genotyped by random amplification of polymorphic DNA (RAPD), pulsed-field gel electrophoresis (PFGE), and flagellin PCR-restriction fragment length polymorphism, and lamb isolates were genotyped by RAPD only. In total, 96% of the poultry and 73% of the lamb samples yielded campylobacters. The lamb isolates were all Campylobacter jejuni, as were 96% of the poultry isolates, with the remainder being Campylobacter lari. The incubation temperature had no significant effect on the number of positive samples or on the species isolated. However, genotyping of the C. jejuni isolates revealed profound differences in the types obtained. Overall (from poultry and lamb), the use of a single incubation temperature, 37 degrees C, gave 56% of the total number of RAPD C. jejuni genotypes, and hence, 44% remained undetected. The effect was especially marked in the poultry samples, where incubation at 37 degrees C gave 47% of the PFGE genotypes but 53% were exclusively recovered after incubation at 42 degrees C. Thus, the incubation temperature of Preston media selects for certain genotypes of C. jejuni, and to detect the widest range, samples should be incubated at both 37 and 42 degrees C. Conversely, genotyping results arising from the use of a single incubation temperature should be interpreted with caution.     

Evaluation of ALOA plating medium for its suitability to recover high pressure-injured Listeria monocytogenes from ground chicken meat

AIMS: To evaluate a chromogenic plating medium for the isolation of sublethally injured cells of Listeria monocytogenes from processed foods. METHODS AND RESULTS: The inactivation of L. monocytogenes at pressures up to 400 MPa and 12 degrees C in ground chicken meat was employed to examine the recovery of high-pressure injured cells. Before and after different repair incubation periods at 30 degrees C in a nonselective broth, samples were plated onto a selective and differential agar [Agar Listeria according to Ottaviani and Agosti (ALOA)] and in the same medium supplemented with 4% sodium chloride (ALOA-S), and incubated at 37 degrees C. Sublethally injured cells were able to grow when directly plated onto the ALOA medium, without a previous repair incubation period. However, only uninjured cells grew on the ALOA-S medium. CONCLUSIONS: Sublethally injured cells of L. monocytogenes can be quantified by subtracting counts on ALOA-S medium from counts on ALOA medium. SIGNIFICANCE AND IMPACT OF THE STUDY: Possible applications include direct enumeration on ALOA of stressed cells of L. monocytogenes in foods with more than 100 colony forming units per gram.     

Effect of temperature, medium composition, and cell passage on production of herpes-based viral vectors

Our work uses replication-defective genomic herpes simplex virus type-1 (HSV-1)-based vectors to transfer therapeutic genes into cells of the central nervous system and other tissues. Obtaining highly purified high-titer vector stocks is one of the major obstacles remaining in the use of these vectors in gene therapy applications. We have examined the effects of temperature and media conditions on the half-life of HSV-1 vectors. The results reveal that HSV stability is 2.5-fold greater at 33 degrees C than at 37 degrees C and is further stabilized at 4 degrees C. Additionally, a significantly higher half-life was measured for the vector in infection culture conditioned serum medium compared to fresh medium with or without serum. Synchronous infections incubated at 33 degrees C produced 2-fold higher amounts of vector than infected cells incubated at 37 degrees C, but with a lag of 16-24 h. Vector production yielded 3-fold higher titers and remained stable at peak levels for a longer period of time in cultures incubated at 33 degrees C than 37 degrees C. A pronounced negative effect of increased cell passage number on vector yield was observed. Vector production at 33 degrees C yielded similar levels regardless of passage number but was reduced at 37 degrees C as passage number increased. Together, these results contribute to improved methods for high-titer HSV vector production.     

Survival in foods of Staphylococcus aureus grown under optimal and stressed conditions and the effect of some food preservatives

Staphylococcus aureus was grown in a rich peptone medium which became alkaline with continued incubation. Cells were grown at 37 degrees C and in the same medium containing 1 M NaCl at 46 degrees C, a temperature at which this organism can grow only when protected by NaCl. Cells of these cultures are hereafter called 37 degrees C-cells and 46 degrees C-cells, respectively. The 37 degrees C-cells harvested when the pH was 7.1 to 7.7 had decimal reduction times (D60-value) of 1.8 to 3.1 min in 50 mM pH 7.2 Tris buffer. The D60 value of 46 degrees C-cells tested in the same way, harvested from cultures at pH 6.6 to 7.6, ranged from 5.3 to a maximum of 12.8 min. In milk, green beans, peas, or beef slurry, the D60-value of 46 degrees C-cells was about four times higher than that of 37 degrees C-cells. Length of survival after freeze-drying in skim-milk powder exposed to air was longest for the cells with the highest D-value. In freeze-dried peas and media acidified with acetic and lactic acids, 46 degrees C-cells survived longer than 37 degrees C-cells. However, the sensitivity of the two kinds of cells to potassium sorbate, sodium benzoate, and sodium propionate was essentially the same, but the 46 degrees C-cells were more resistant to butylated hydroxyanisole and sodium nitrite.     

Effects of sample handling temperatures on bovine skim milk progesterone concentrations

The effects of incubation of whole milk at various temperatures and times on the concentration of progesterone in the skim milk fraction was determined. For the study, milk samples were collected from 10 pregnant Holstein cows. The milk from each cow was transferred to culture tubes to provide 32 replicates of 3 ml volume. To begin the incubation study, all samples were placed in a 37 degrees C water bath for 4 h. The end of this incubation was designated as time 0 and a sample from each cow was centrifuged to harvest skim milk. At time 0, samples from each cow were divided among incubation temperatures of 0 degrees, 4 degrees, 20 degrees and 37 degrees C. Samples were removed from each incubation group at 30, 60, 90 and 120 min. After 120 min, all remaining samples were returned to the 37 degrees C incubation and skim milk was collected at 30, 60 and 90 min. Progesterone was measured in skim milk by radioimmunoassay. The mean +/- SE concentration of progesterone in skim milk at time 0 was 10.9 +/- 1.1 nmol/L. The mean concentration of progesterone in skim milk was higher (P < 0.05) in all samples incubated at 0 degrees and 4 degrees C, with incremental increases ranging from 34% to 67% above time 0. Progesterone in skim milk returned to time 0 concentrations in milk samples transferred from 0 degrees or 4 degrees C to 37 degrees C. There was no change in skim milk progesterone in whole milk samples incubated at 20 degrees or 37 degrees C. From this study, it can be concluded that the concentration of progesterone in skim milk is temperature dependent. Inconsistency in handling whole milk samples can have a profound effect in the concentration of progesterone on skim milk. The temperature-dependent effect was reversible and may be related to solubility of progesterone in milk fat.     

Comparison of Methods for Isolation of Mycobacteria from Water

Twelve methods for the isolation of mycobacteria were compared by applying them in parallel to 26 samples of surface water and 109 samples of treated water. Each method was defined by a particular combination of decontamination method, growth medium, and incubation temperature. For the decontamination of surface water, we used cetylpyridinium chloride (CPC) (30 min, 0.05%), as well as sample preincubation in tryptic soy broth (TSB) followed by decontamination with a cocktail of NaOH, cycloheximide, and malachite green. Treated water was decontaminated with 0.005 and 0.05% CPC (30 min). After enrichment by filtration, all samples were incubated on Lowenstein-Jensen medium (LJ), Ogawa egg yolk medium (OEY), and Ogawa whole-egg medium containing ofloxacin and ethambutol (OEOE) at temperatures of 30 and 37(deg)C. The efficacy of each method was determined by calculating the positivity rate, negativity rate, contamination rate, mean number of mycobacterial colonies grown, and mean number of different mycobacterial strains isolated. The last value was determined by subjecting the isolates to PCR restriction analysis and mycolic acid thin-layer chromatography. Statistical analysis demonstrated that both the TSB method and 0.05% CPC were appropriate for the decontamination of surface water. Decontamination with 0.005% CPC was best for treated water. The results for incubation on LJ were at least equal to those for incubation on OEY and always superior to the results with OEOE. At an incubation temperature of 30(deg)C, all methods achieved higher yields than at 37(deg)C.     

Muscle cell cultures from chicken breast muscle have increased specific activities of creatine kinase when incubated at 41 degrees C compared with 37 degrees C

Chicken muscle cell cultures were incubated at 41 degrees C, the physiological chicken body temperature, and compared with cultures incubated at 37 degrees C, the typical cell culture incubation temperature. The cultures incubated at 41 degrees C show not only an increase in creatine kinase (CK)-specific activity but also a marked increase in the percentage of adult muscle CK isozyme (MM-CK) in 7-day muscle cultures. Muscle cell cultures incubated in the presence of cytosine arabinoside (ara-C), a cell proliferation inhibitor, do not have the mononucleated cell overgrowth seen at 41 degrees C and thus exhibit a further increase in creatine kinase-specific activity compared with cultures incubated at 41 degrees C in the absence of ara-C. These results suggest that muscle cell cultures incubated at 41 degrees C are more highly differentiated than those incubated at 37 degrees C.     

Separation of the epithelial and mesenchymal components of the newt limb regenerate with salt.

After incubation of isolated forelimb regenerates of Notophthalmus (Triturus) viridescens at all developmental stages for 60 minutes at 37 degrees C in a salt medium containing 111 mM sodium chloride, 5.6 mM potassium chloride and 100 mM sodium phosphate buffer at pH 7.5, the wound epithelium of each regenerate was removed intact from its underlying mesenchymal component. The suggestion is made that the salt medium is an effective epithelial-mesenchymal separating agent due to a combination of its hypertonicity, high ionic strength and the fact that the medium precipitates calcium as calcium phosphate. Attempts to dissect away the epithelium from the mesenchyme after incubation of isolated regenerates in sodium phosphate containing 1% or 3% Difco 1:250 trypsin, 10 mM EDTA or 150 units collagenase/ml medium were unsuccessful. Epidermis of adult newt forelimb skin was removed only after extended incubation of the forelimbs in the salt medium for three hours at 37 degrees C or after freezing isolated forelimbs in buffer and subsequent thawing.     

Externalization of transferrin receptor in established human cell lines

The externalization of transferrin receptors was found in established human tumor cell lines at the rate of 10-35 ng/hour/10(6) cells, when they were incubated with transferrin at 37 degrees C. This externalization is inhibited by lowering the incubation temperature to 4 degrees C or eliminating the ligand from the culture medium. Metabolic inhibitors such as sodium azide, colchicine, cytochalasin B and chloroquine also decreased the rate of externalization. Almost 95% of released transferrin receptors were precipitated by centrifugation at 100,000 x g for 30 min, suggesting that transferrin receptor is externalized into the medium as a vesicular form.     

Effect of caffeine on the post-thaw motility of buffalo spermatozoa

Buffalo semen was diluted (1:2) with lactose diluent containing caffeine (2, 4 and 6 mM). Diluted semen samples were frozen in a pellet form (0.15 ml), thawed 24 h after freezing in 2.9% sodium citrate for 30 sec and incubated at 37 degrees C for 3 h. Addition of caffeine to diluted buffalo semen before freezing resulted in a significant increase in the post-thaw motility of spermatozoa over the 3-h incubation period. When caffeine was added to the thawing medium, the post-thaw motility was further improved. Thus, the increase in motility due to caffiene treatment was even more pronounced than in samples treated with caffiene before freezing.     

Interactive effects of solutes, potassium sorbate and incubation temperature on growth, heat resistance and tolerance to freezing of Zygosaccharomyces rouxii.

The interactive effects of solutes, potassium sorbate and incubation temperature on growth, heat resistance and tolerance to freezing of Zygosaccharomyces rouxii were investigated. Growth rates in media supplemented with glucose, sucrose or NaCl to aw 0.93 were more rapid than in unsupplemented media (aw 0.99). Although growth in unsupplemented medium was lower at 35 degrees C, incubation at 21 degrees C or 35 degrees C had little effect on growth in media supplemented with glucose and sucrose. The addition of 300 micrograms potassium sorbate/ml to media resulted in reduced growth rates, particularly at 35 degrees C. Heat resistance of Z. rouxii was substantially greater in cultures previously incubated at 35 degrees C than in cultures incubated at 21 degrees C in media both with and without 300 micrograms potassium sorbate/ml. Zygosaccharomyces rouxii was tolerant to freezing at -18 degrees C for up to 120 d in all test media supplemented with glucose, sucrose or NaCl. The addition of 300 micrograms potassium sorbate/ml to sucrose-supplemented media resulted in increased resistance to freezing in cultures previously incubated at 21 degrees C. Sensitivity to freezing increased when cultures were incubated at 21 degrees C in media not supplemented with solutes. Glucose and sucrose provided the best protection against inactivation by heating and freezing, regardless of the presence of potassium sorbate in growth media.     

Influence of the sporulation temperature upon the heat resistance of Bacillus subtilis.

The influence of different sporulation temperatures (30, 37, 44 and 52 degrees C) upon heat resistance of Bacillus subtilis was investigated. Heat resistance was greater after higher sporulation temperatures. Relation of heat resistance and temperature of sporulation was not linear over all the range of temperatures tested. Heat resistance increased about tenfold in the range of 30-44 degrees C. Sporulation at 52 degrees C did not show any further increase in heat resistance. This effect was constant over all the range of heating temperatures tested (100-120 degrees C). z value remained constant (z = 9 degrees C). Greater heat resistances at higher temperatures of sporulation were not due to selection of more heat resistant cells by a higher sporulation temperature. Spores obtained from cells incubated at 32 or 52 degrees C always possessed heat resistances that corresponded to the sporulation temperature regardless of the incubation temperature of their vegetative cells.     

Influences of incubation temperature and various saccharides on the production of organic acids and gases by gut microbes of rainbow trout Oncorhynchus mykiss in a micro-scale batch culture

We studied the influence of incubation temperature and additional saccharides on the metabolism of hindgut microbes of the rainbow trout Oncorhynchus mykiss in a 50 microl-scale batch culture system. Intestinal contents of rainbow trout reared at 15 degrees C were incubated with glucose, lactosucrose, sodium alginate or colloidal chitin (each 10 g/l) at 15 degrees C or 25 degrees C for 12 h. Levels of organic acids at 0 h and 12 h of incubation were quantified with HPLC. We also monitored gas release from these cultures during incubation. The main product was iso-butyric acid, except for the cultures with colloidal chitin where no net production of organic acids was observed. We detected higher levels of iso-butyric acid in cultures with lactosucrose than in the other cultures. Net production of this acid was less in cultures with colloidal chitin than in blank cultures. The volume of released gas was larger when incubated at 25 degrees C than at 15 degrees C. Cultures with colloidal chitin released more gas than blank cultures when they were incubated at 15 degrees C. Cultures with sodium alginate released less gas than blank cultures irrespective of incubation temperature. These results indicate that the hindgut microbes of this carnivorous fish mainly produce branched-chain fatty acids, very likely by microbial digestion of nitrogenous materials rather than saccharides. However, additional saccharides affected production of branched-chain fatty acids. The influence of incubation temperature in the present study also suggested that the environmental temperature of host fish should affect microbial digestion in the fish gut.     

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.     

Binding, internalization, and lysosomal association of 125I-human growth hormone in cultured human lymphocytes: a quantitative morphological and biochemical study

125I-human growth hormone (125I-hGH) binds specifically to receptors on cultures human lymphocytes (IM-9). When this process is studied by use of quantitative EM radioautography, under conditions of incubation at 15 degrees C for 5 min, the ligand is localized to the plasma membrane of the cell. At 30 degrees and 37 degrees C, however, 125I-hGH is progressively internalized by the cell as a function of time. The internalized ligand is found predominantly in the Golgi region of the cells, with a five-fold preferential localization to membrane-bounded structures with the morphological and cytochemical characteristics of lysosomes. Up to 59% of these lysosome-like structures are positive for the acid phosphatase reaction under the conditions of incubation at 37 degrees C for 120 min. When the cell associated radioactivity after 15- 120 min of incubation at 37 degrees C is extracted in 1 M acetic acid and filtered on a Sephadex G-100 column, 58-73% of the material elutes as intact hGH. When cells are incubated with 125I-hGH at 37 degrees C for 15-120 min, separated from the incubation medium, and washed and diluted 100-fold, the percent 125I-hGH dissociable decreases as a function of increasing time of incubation. When cells are incubated with 125I-hGH for 15 min at 37 degrees C and the radioactivity that dissociates from the cells during 15-90 min is studied, the labeled material appearing in the incubation medium is progressively degraded as a function of time of incubation. When the dissociation process is studied radioautographically, grains are found both in plasma membrane and intracelluar compartments after 30 min of association, but after 30 and 120 min of dissociation a higher proportion of grains are in the intracellular compartment. After 120 min of association, there is less dissociation from either compartment and a preferential increase of grains in the intracellular compartment. These data suggest that receptor-linked internalization of a polypeptide hormone provides a mechanism that couples degradation of the ligand with loss of the cell surface receptor.     

Comparison of the Effects of Liquid Medium Repair and the Incorporation of Catalase in MacConkey Type Media on the Recovery of Enterobacteriaceae Sublethally Stressed by Freezing

Nine pure cultures of species of Enterobacteriaceae were stressed by rapid freezing in tryptone soya broth (TSB) to — 22°C and subsequent storage at that temperature for 7 d. About one to two log cycles kill and at least one additional log cycle sublethal impairment was achieved. Numbers of colonies of these cultures in poured plates of violet red bile glucose (VRBG) agar, with 67 u/ml of catalase added at 47°C, were only slightly higher than those in plain VRBG, both incubated overnight at 30°C. Two hours incubation of TSB suspensions at 17–25° C resulted in almost complete restoration of the ability of cells to develop colonies in VRBG, without, however, leading to any significant multiplication.
Similar experiments with 32 samples of frozen minced meat, 27 samples of frozen surface water, 18 of frozen chicken liver and 14 of fresh sausage substantiated the results obtained in the studies on pure cultures.
In the experiments with the nine pure cultures the influence of the nutrient composition of the solid enumeration media: 'minimal' agar, TSB agar (TSBA) and Mueller-Hinton agar with Polyvitex nutrient supplement (MHA), on the recovery of Enterobacteriaceae stressed by freezing was also studied. Colony numbers in TSBA and MHA were virtually identical. The glucose mineral salts medium led to lower recovery, indicating that so-called 'minimal medium recovery' of stressed bacterial populations is not a common phenomenon.     

Effect of osmotic pressure on neurogenesis in cultures of chich embryo spinal cords.

The osmotic pressure of the medium in stoppered, roller tube cultures increased by an average of 17 +/- 6 mOsM per kg of water during 3 days of incubation at 37 degrees C irrespective of the initial osmolality (280 to 340 mOsM) of the medium. The increase was apparently due to evaporation of water from the medium into the gas phase of the roller tube. This observation led us to study the effect of osmotic pressure on neuronal differentiation in cultures of chick embryo spinal cords. Spinal cords were excised from stage 16 to 19 (2.5 to 3 days of incubation) or stage 36 (10 days) chick embryos and cultured as fragments on collagen-coated cover slips in roller tubes at 37 degrees C for 21 days. The medium was adjusted to 283 +/- 3,300 +/- 3,323 +/- 3, or 342 +/- 3 mOsM per kg with saturated choline chloride solution or distilled water. The results indicate that the nature of the neuronal differentiation in vitro was not altered by the osmolality of the medium. The proportion of cultures containing neurons was influenced by osmolality. In the 300 +/- 3 mOsM medium, 75% of all the stage 36 cultures initiated contained neurons, and 52% of all the stage 16 to 19 cultures initiated contained neurons. In the other media the proportion of neuron-containing cultures was lower. Two conclusions were drawn. Neurogenesis in cultures of embryonic chick spinal cord fragments is sensitive to an increase in the initial osmotic pressure of the medium as small as 20 mOsM above the optimal 300 mOsM. As a result of the 17 mOsM increase which always occurred in the culture medium between feedings, the optimum osmolality for neuronal development is in fact a range, from 300 to 317 mOsM.     

Changes in bleb formation following hyperthermia treatment of Chinese hamster ovary cells

Chinese hamster ovary cells in suspension cultures were heated for various times at 41.5, 43.5, and 45.5 degrees C, and quantitative determinations of microblebbing and macroblebbing of the cell membrane were performed for cells maintained at 4, 25, and 37 degrees C after hyperthermia. The percentage of cells with blebs following heating at 45.5 degrees C was dependent upon the duration of heating with increases from 40% for 5 min to 90% for 30 min. Cells exposed to lower temperatures exhibited less blebbing which was not quantifiable. The changes in bleb formation following 45.5 degrees C were dependent upon the posthyperthermia temperature: a slight decrease of macroblebbing at 25 degrees C, a decrease to 50% by 2 h at 37 degrees C, and a sharp decrease of macroblebbing to less than 10% by 1 h at 4 degrees C. Microblebbing increased slightly at 37 degrees C. When cells were transferred rapidly from the 4 degrees C posthyperthermia incubation to 37 degrees C, the bleb formation percentages returned rapidly to the higher levels which existed before posthyperthermia incubation at the lower temperatures. Gamma irradiation of 20 and 50 Gy produced only a small increase in microblebbing at longer periods (5 to 6 h) but no increase in macroblebbing. The survival of cells heated for 20 min at 45.5 degrees C was decreased 40% for suspension cells maintained at 4 degrees C for 2 to 3 h before incubation at 37 degrees C for colony formation compared to cells immediately incubated at 37 degrees C after heating. The survival of cells maintained at 25 degrees C after heating was not altered in comparison.     

Mechanisms of loss of latency of lysosomal enzymes. Effects of incubation on the properties of lysosomal membranes.

1. The effects of sucrose and KCl on the loss of latency of lysosomal enzymes caused by incubation at 37 degrees C, pH 7.4, were examined by using Triton-filled lysosomes from rat liver and two fractions from livers of rats not injected with Triton. 2. After incubation, the percentage free activity of lysosomal enzymes was measured before and after cooling to 0 degrees C in order to determine the amount of latency lost at 37 degrees C without cooling and the additional amount lost on cooling the incubated lysosomes to 0 degrees C. 3. The latency that is lost without cooling is first decreased and then increased by increasing the osmotic strength of the incubation medium with KCl, or with sucrose in the presence of KCl. However, if the osmotic strength is increased with sucrose alone, loss of latency is decreased up to 0.25M-sucrose, but is increased only slightly at higher sucrose concentrations. Apparently the lysosome is permeated by hyperosmolar KCl but not by sucrose during incubation. 4. If the osmotic strength of the assay medium is increased with KCl, the loss of latency caused by incubation for 60 min in hyperosmolar KCl is repressed. Thus it appears that a KCl-permeated lysosome can be obtained which is relatively stable until exposure to lower osmolarities. 5. The loss of latency caused by cooling incubated lysosomes to 0 degrees C is largely eliminated if the osmotic strength of the medium in which the lysosomes are cooled is raised sufficiently with either sucrose or KCl. 6. Osmotic-fragility curves were obtained after incubation for 1 and 60 min at iso-osmoticity (0.2M-KCl or 0.25 M-sucrose). Although little loss of latency occurs at iso-osmoticity, lysosomes incubated for 60 min display greatly increased fragility on exposure to hypo-osmolar KCl, hypo-osmolar sucrose or hyperosmolar KCl. 7. It is suggested that permeability to KCl at 37 degrees C and the increase in fragility on exposure to hypo-osmolar conditions are both consequences of injury, probably from enzymic action, sustained by the lysosomal membrane during incubation at 37 degrees C.     

The effect of supraoptimal temperature on the multiplication of different cytomegalovirus strains.

The virion synthesis by five human cytomegalovirus (CMV) strains in human embryonic fibroblast cultures was stopped by incubation of the infected cultures at 40 degrees C. At this temperature the antigens appeared diffusely filling the nucleus and the cytoplasm. The blocking effect of the elevated temperature was exerted in the same period of the reproduction cycle as the inhibitory effect of cytosine arabinoside (ara-C). In cell cultures infected with CMV and incubated first at 40 degrees C, then at 37 degrees C, the synthesis of infectious virus started again, thus the abortive cycle developed at 40 degrees C was reversible. The inhibition of virus multiplication cannot be attributed to the thermosensitive events in the normal function of the host cell.