Effect of Relative Humidity and Temperature on Airborne Venezuelan Equine Encephalitis Virus

Inactivation of airborne Venezuelan equine encephalitis (VEE) virus disseminated from liquid suspensions or from lyophilized preparations as 1- to 5-mum particles was investigated under various conditions of relative humidity and temperature in a 2,500-liter static aerosol chamber. Relative humidity ranging from 18 to 90% at 24 C and temperature ranging from -40 to 24 C had no marked effect on the biological decay rate or the recovery of viable airborne VEE virus disseminated from liquid suspensions. However, at 49 C a significant increase in the biological decay rate and decrease in aerosol recovery of the VEE virus were observed. Airborne lyophilized VEE virus was significantly affected by relative humidity. An increase in relative humidity from 20 to 90% resulted in progressive decrease in aerosol recovery of viable VEE virus. A twofold reduction in aerosol recovery of the lyophilized virus was observed at and above 29 C as compared to the lower temperatures studied. However, the differences among biological decay rates of lycphilized VEE virus were not significant within temperature range of -40 to 38 C.     

Relationship Between Atmospheric Temperature and Survival of Airborne Bacteria

Effects of temperatures ranging from -40 to 49 C on the behavior of airborne Serratia marcescens, Escherichia coli, and Bacillus subtilis var. niger were investigated. Aerosol decay rates of B. subtilis spores were not significantly affected by the temperature and remained approximately constant within the temperature range studied. The survival of airborne S. marcescens and E. coli was closely related to the temperature. An increase in temperature from -18 to 49 C resulted in a progressive increase of the biological death rate, and the relationship between the biological death rate and the temperature appeared to be linear. An increase in temperature from 24 to 49 C resulted in significantly reduced aerosol recoveries of the two vegetative organisms. At -40 C, the aerosol recovery of all three agents was consistently lower than at -18 to 24 C.     

Effects of Atmospheric Humidity and Temperature on the Survival of Airborne Flavobacterium

The survival of airborne Flavobacterium sp. in particle sizes ranging from 1 to 5 μm was significantly influenced by atmospheric temperature. A progressive increase in temperature from −18 to 49 C resulted in increases in death rates of the airborne organism. The lowest death rates were observed in the temperature range of −40 to −18 C, and the highest death rates were observed in the 29 to 49 C range. At 24 C, the survival of airborne Flavobacterium did not appear to be significantly affected by relative humidity ranging from 25 to 99%.     

Effect of NO2 on Airborne Venezuelan Equine Encephalomyelitis Virus

Studies were conducted to determine the effect of nitrogen dioxide (NO(2)) on aerosol survival and biological decay rate of Venezulean equine encephalomyelitis (VEE) virus and spores of Bacillus subtilis var. niger. The NO(2) concentrations used in the experiments were 0.5, 5, and 10 ppm at 24 C and 85% RH. The survival of airborne VEE virus disseminated as particles 1 to 5 mum in diameter was significantly influenced by the presence of 5 ppm of NO(2). At this concentration, the biological decay rate increased threefold and the aerosol recovery and aerosol survival of the VEE virus were significantly lower than at 0.5 ppm or in the absence of NO(2). Airborne spores of B. subtilis were not significantly affected by as much as 10 ppm of NO(2).     

Effects of Environmental Factors on the Survival of Airborne T-3 Coliphage

Experiments were conducted to determine the effects of storage temperatures, relative humidity, and additives on the survival of aerosolized Escherichia coli phage T-3. The aerosol stability of the coliphage, calculated as per cent recovery, was not affected by storage at 10 or -70 C for up to 4 months. However, an increase in aerosol decay rate of coliphage stored at 10 C was observed. The effect of humidities ranging from 20 to 90% relative humidity was studied, and it was observed that humidities lower than 70% relative humidity significantly reduce the survival of airborne coliphage. The effect of various compounds on the aerosol decay rate of T-3 coliphage was studied at 50 and 85% relative humidity. Addition of dextrose in 0.1 M concentrations to the disseminating fluid significantly reduced aerosol decay rate at 50% relative humidity without affecting the decay at 85%. Addition of spermine, spermidine-phosphate, thiourea, galacturonic acid, and glucosaminic acid, individually or in combination, had no effect on aerosol decay rates. The use of deuterium oxide as the suspending fluid for dissemination had no effect on aerosol stability of the coliphage.     

Effect of epididymis handling conditions on the quality of ram spermatozoa recovered post-mortem

Post-mortem spermatozoa recovery is an important technique for obtaining germplasm reserves from genetically valuable animals or endangered species. However, there are many factors that influence the outcome of this technique. We have studied the effect of the interval between animal's death and sperm recovery (0, 24 or 48 h) on the quality and freezability of ram spermatozoa from cauda epididymidis. Storage temperature of epididymis (room temperature or 5 degrees C) was also analysed. Spermatozoa were diluted with Tes-Tris-Fructose solution supplemented with egg yolk (10%) and glycerol (4%), and frozen using a programmable biofreezer (-20 degrees C/min). Pre-freeze and post-thaw sperm samples showed viable spermatozoa up to 48 h after the animal's death, although their quality declined significantly as post-mortem storage time increased. Epididymis sperm stored at 5 degrees C showed better motility and a lower percentage of abnormal forms than epididymis stored at room temperature after 24 and 48 h. The fertilizing ability of cauda epididymis ram spermatozoa obtained at 0 and 24h after the animal's death is similar to that of ejaculated spermatozoa. Therefore, a good protocol for post-mortem semen collection in rams when epididymal spermatozoa cannot be collected immediately, is to preserve the epididymis at 5 degrees C and process the samples in the first 24h after the animal's death.     

Survival of Airborne Bacteria in a High Urban Concentration of Carbon Monoxide

Vegetative cells of Serratia marcescens 8UK, Sarcina lutea, and spores of Bacillus subtilus var. niger were held in aerosols, with and without an urban concentration of CO (85 muliters per liter or ppm), for up to 6 hr at 15 C and a relative humidity (RH) of approximately 0, 25, 50, 75, and 95%. It was found that CO enhanced the death rate of S. marcescens 8UK at least four- to sevenfold at low RH (ca. 1 to 25%), but protected the cells at high RH (ca. 90%). Death rates of S. lutea, with or without added CO, were comparatively low over the entire RH range. However, in the first hour, airborne S. lutea held in CO-containing air were more stable than those in air without added CO (i.e., CO protection). A marked increase in the death rate (up to 70-fold) occurred in the subsequent 5 hr within the RH range of approximately 0 to 75%. Statistical analysis indicated that aerosol decay rates of B. subtilus var. niger spores decreased significantly, when held in a CO-containing as compared to a non-CO-containing atmosphere, in the 0 to 85% RH range. Thus, the data presented indicate that CO in the urban environment may have a protective or lethal effect on airborne bacteria, dependent upon at least the microbial species, aerosol age, and relative humidity. A mechanism for CO death enhancement and protection of airborne S. marcescens 8UK is suggested to involve CO uncoupling of an energy-requiring death mechanism and an energy-requiring maintenance mechanism at high and low RH, respectively.     

Survival of Bacteria on Metal Surfaces

Survivor curves were determined for Serratia marcescens, Sarcina lutea, Pasteurella tularensis, and P. pestis deposited from the airborne state onto metallic surfaces and subsequently stored at various humidities and temperatures. Cells of all species tested remained alive longest in a dry atmosphere, except that cells of S. marcescens survived best in a saturated atmosphere. Survival decreased most rapidly at the intermediate humidity level for three of the test organisms, yet P. tularensis died most rapidly in a saturated atmosphere. An increase in temperature decreased survival of P. pestis and P. tularensis.     

A statistical model of laboratory death rate measurements for airborne bacteria

Summary From 270 published laboratory airborne death rate measurements, two regression models relating the death rate constant for 15 bacterial species to aerosol age in the dark, Gram reaction, temperature, and an evaporation factor which is a function of RH and temperature were obtained. The independent variables accounted for 94% of the variation in the data for each of the two models. In both models the regression shows an increased survival rate with aerosol age accounting for approximately 90% of the total variation in the data. The remainder of the total variation was explained by temperature and RH (in interaction with the Gram reaction) in one model and by the evaporation function (in interaction with the Gram reaction) in the order model. Death rate data for gaseous atmospheric contamination, and light experiments were too few for building a regression model. In addition, these points were not well fit by the model indicating further research is needed to prepare realistic prediction models for airborne bacterial survival.     

Airborne Stability of Tailless Bacterial Viruses S-13 and MS-2

The effect of relative humidity (RH) on the airborne stability of two small bacterial viruses, S-13 and MS-2, was studied. Poorest recovery of S-13 was obtained at 50% RH. Humidification prior to aerosol sampling significantly increased the recovery of S-13 at RH deleterious to the airborne virus. A commercial preparation of MS-2 suspended in a buffered saline solution showed a rapid loss of viability at RH above 30%, whereas a laboratory preparation containing 1.3% tryptone showed high recoveries at all RH studied. Dilution of the commercial MS-2 into tryptone broth conferred stability on the airborne virus. Humidification prior to sampling significantly reduced the viable recovery from aerosols of commercial MS-2, whereas the laboratory preparation was unaffected.     

Enhanced Recovery of Airborne T3 Coliphage and Pasteurella pestis Bacteriophage by Means of a Presampling Humidification Technique

This paper reports a series of experiments in which two methods of collecting airborne bacteriophage particles were compared. A standard aerosol sampler, the AGI-30, was evaluated for its competence in measuring the content of bacteriophage aerosols. It was used alone or with a prewetting or humidification device (humidifier bulb) to recover T(3) coliphage and Pasteurella pestis bacteriophage particles from aerosols maintained at 21 C and varied relative humidity. Collection of bacteriophage particles via the humidifier bulb altered both the initial recovery level and the apparent biological decay. Sampling airborne bacteriophage particles by the AGI-30 alone yielded data that apparently underestimated the maximal number of potentially viable particles within the aerosol, sometimes by as much as 3 logs.     

Improved Procedure for Identification of Group D Enterococci with Two New Media

The aerosol survival in air was determined for Pasteurella tularensis live vaccine strain (LVS) as a function of relative humidity (RH). Three different preparations of bacteria were used: (i) liquid suspension of P. tularensis LVS in spent culture medium; (ii) powders of P. tularensis LVS freeze-dried in spent culture fluid; (iii) P. tularensis LVS freeze-dried in spent culture fluid and then reconstituted with distilled water and disseminated as a liquid suspension. Preparation (i) gave greatest survival at high RH and lowest survival at intermediate RH. Preparation (ii), in contrast, gave greatest survival at low RH and minimum survival at 81% RH. Preparation (iii) was the same as preparation (i), i.e., the process of freeze-drying and reconstituting with distilled water before aerosol formation had little or no effect upon aerosol survival as a function of RH. Hence, control of aerosol survival appears to be through the water content of P. tularensis LVS at the moment of aerosol generation rather than the water content of the bacteria in the aerosol phase.     

The effect of aerosolization on the changes in major biological properties of the vaccine strain of Francisella tularensis

The exposure of F. tularensis vaccine strain 15/10 in the form of aerosol for more than 10 minutes results in the decrease of its virulence, immunogenicity and the content of species-specific antigen and in the increase of the dissociation level. Proceeding from this fact, during aerosol immunization with this vaccine strain exposure must not exceed 5 minutes for unstabilized aerosol and 10 minutes for aerosol stabilized with 5% glycerin. Under these conditions the properties of F. tularensis strain 15/10 are retained practically on the initial level.     

PCR和Southern Blot检测土拉弗氏菌气溶胶

为提高检测土拉弗氏菌的特异性和敏感性,建立了土拉菌ＰＣＲ及核酸杂交检测方法。运用平板计数、多聚酶链反应对土拉菌气溶胶稳定性进行了比较,结果表明ＰＣＲ具有较高灵敏度,并且在采样后３小时ＰＣＲ就可以得出定性结果,而平板计数则需要３～７天。采用ＰＣＲ法合成了土拉菌３７６－ｂｐ探针,分别对细菌菌液、５６８－ｂｐＰＣＲ产物和气溶胶样品进行杂交,结果表明菌悬液直接杂交可检出１０５ＣＦＵ左右的细菌,检测ＰＣＲ产物可达４０ｐｇ。ＰＣＲ和Ｓｏｕｔｈｅｒｎ印迹相结合有利于细菌的分离鉴定     

The effects of low temperature on fertilized rabbit ova in vitro,and the normal development of ova kept at low temperature for several days

Fertilized rabbit ova at the 2-blastomere stage kept in rabbit serum were stored at low temperatures for various lengths of time. They were then cultured at 38 degrees C. for about 24 hours to determine their viability. A number of the viable ova were finally transplanted into recipient does. It was found that rapid cooling of ova to 5 degrees or to 0 degrees C. was more harmful to the subsequent viability of ova than slow cooling. Rapid cooling was not more lethal to the ova than slow cooling, but did prevent their future normal cleavage. There was no difference between those ova cooled rapidly or slowly to 10 degrees C. It was concluded that temperature shock has an adverse effect on ova, especially at the lower temperatures, though temperature shock can be remedied by acclimatization (slow cooling). Thus, the physiological significance of temperature shock would seem to be broadened. The optimal temperature for the storage of ova was investigated. It was found that 10 degrees C. was the best temperature; at this temperature viable ova were obtained after storage for 144 to 168 hours. At 0 degrees , 5 degrees , or 15 degrees C. the ova were viable for 96 to 120 hours, while at 22-24 degrees C., only for 24 to 48 hours. The percentage of dead ova was low at a favorable temperature, increasing only at the end of the storage period. At an unfavorable temperature, however, the rate of death increased steadily from beginning to end of storage. The percentage of abnormally cleaved ova (arrested cleavage and fragmentation) remained at a low level at first at a favorable temperature, but then increased just before or during death of the ova. A critical time for the viability, the abnormal cleavage, and the death of ova was characteristic of each temperature. About 24 to 28 per cent of the viable ova remaining after being stored at 0-15 degrees C. for 2 to 4 days and cultured at 38 degrees C. for 24 hours were capable of development into normal young. The compatibility of serum and ova, the absence of a correlation between the viability of the ova and the source of the fertilizing spermatozoa, and the fertilization of superovulated ova (i.e., the percentage of fertile does in follicular phase and in luteal phase, the percentage of unfertilized ova and of fertilized ova at different stages, the percentage of does that had produced a normal number of ova or had produced a large number of ova, etc.), are reported. The possibility of a more efficient utilization of the germ cells of valuable animals by means of the present techniques, and the possibility of a new approach to the experimental investigation of mammalian genetics and development, have been mentioned.     

Toxicity of Pasteurella tularensis killed by ionizing radiation

Approximately 2 x 10(11) viable Pasteurella tularensis cells per ml, contained in suspensions, were killed by exposure to 10(6) r of gamma-radiation. When injected intraperitoneally into mice, the irradiated suspensions initially contained about 10 ld(50) per ml, and immunized mice against challenge with fully virulent strains of P. tularensis. Toxicity and immunizing activity of the suspensions decreased significantly within a few days at 5 C. Mice were protected against the toxin by immune serum or by prior injection of endotoxin of Escherichia coli. Cortisone did not protect against the newly prepared suspension, but was effective against the aged suspension. Lethal doses of newly prepared suspension for guinea pigs and rabbits were approximately 0.5 ml and 2 ml, respectively. Cortisone protected rabbits, but not guinea pigs, against lethal challenge. Pyrogenic effects resembling those shown by endotoxin-containing suspensions were demonstrated in rabbits. The results suggested that two toxins are responsible for the toxicity of irradiated suspensions of P. tularensis: one labile and associated with the immunizing activity of the suspension, the other more stable and resembling classical endotoxin.     

Induced encystment improves resistance to preservation and storage of Acanthamoeba castellanii

Several conditions that allow the preservation, storage and rapid, efficient recovery of viable Acanthamoeba castellanii organisms were investigated. The viability of trophozoites (as determined by time to confluence) significantly declined over a period of 12 months when stored at -70 degrees C using dimethyl sulfoxide (DMSO; 5 or 10%) as cryopreservant. As A. castellanii are naturally capable of encystment, studies were undertaken to determine whether induced encystment might improve the viability of organisms under a number of storage conditions. A. castellanii cysts stored in the presence of Mg2+ at 4 degrees C remained viable over the study period, although time to confluence was increased from approximately 8 days to approximately 24 days over the 12-month period. Storage of cysts at -70 degrees C with DMSO (5 or 10%) or 40% glycerol, but not 80% glycerol as cryopreservants increased their viability over the 12-month study period compared with those stored at room temperature. Continued presence of Mg2+ in medium during storage had no adverse effects and generally improved recovery of viable organisms. The present study demonstrates that A. castellanii can be stored as a non-multiplicative form inexpensively, without a need for cryopreservation, for at least 12 months, but viability is increased by storage at -70 degrees C.     

In vitro interactions between rabbit alveolar macrophages and Pasteurella tularensis

Nutter, J. E. (Fort Detrick, Frederick, Md.), and Q. N. Myrvik. In vitro interactions between rabbit alveolar macrophages and Pasteurella tularensis. J. Bacteriol. 92:645-651. 1966.-Rabbit alveolar macrophages were successfully employed in a study of host cell-Pasteurella tularensis interactions in vitro. Under cell culture conditions in which inhibitory antibiotics were not employed and small infection ratios were used, the relative in vivo virulence of two strains of P. tularensis was duplicated. As a consequence of intracellular multiplication, normal macrophages were killed in relation to the virulence of the strain employed. Alveolar macrophages were also collected from immune rabbits, and macrophage mortality and bacterial growth were significantly suppressed below levels observed with normal macrophage preparations. The effect of immune serum could only be ascribed a minor role in the observed reactions. A marked intravenous toxicity of P. tularensis for the rabbit was observed with both virulent and attenuated strains. The toxicity was possessed only by viable preparations and could be elicited in animals immune to virulent challenge.     

Reproductive rates of Russian wheat aphid (Hemiptera: Aphididae) biotypes 1 and 2 on a susceptible and a resistant wheat at three temperature regimes

The reproductive rates of Russian wheat aphid, Diuraphis noxia (Kurdjumov) (Hemiptera: Aphididae), Biotype 1 (RWA 1) and Biotype 2 (RWA 2) were compared in the laboratory at three temperature regimes on a Russian wheat aphid resistant cultivar ('Prairie Red') and a susceptible cultivar ('TAM 107'). The objective of this study was to expose RWA 1 and RWA 2 to three temperature regimes and two levels of resistance to find whether there were reproductive differences that may occur within each biotype as well as differences in reproduction between biotypes. In addition, temperature effects of the Dn4 gene on biotype reproduction were noted. Differences in reproductive rates between the two biotypes seem to be driven by temperature. For both biotypes, longevity and reproductive rate parameters, except for intrinsic rate of increase, were lower at the 24-29 degree C temperature regime than the 13-18 degree C and 18-24 degree C temperature regimes. The intrinsic rate of increase was higher for both biotypes at the 18-24 degree C and 24-29 degree C temperature regimes than at the 13-18 degree C temperature regime. Reproductive rates between biotypes were similar at the two higher temperature regimes, but the fecundity for RWA 1 was less than RWA 2 at the 13-18 degree C temperature. The change in fecundity rates between RWA 1 and RWA 2 at lower temperatures could have ecological and geographical implications for RWA 2.     

Influence of endophyte-infected tall fescue on cyclicity, pregnancy rate and early embryonic loss in the mare

The effects of grazing endophyte-infected tall fescue on luteal function, pregnancy rates, and embryonic loss rates were compared between treated mares (n=18) and untreated controls (endophyte-free, n=12). Mares grazing endophyte-infected fescue demonstrated significantly (P<0.01) prolonged luteal function (22.9 vs 15.8 d) than those grazing endophyte-free fescue. Continuous grazing of endophyte-infected fescue resulted in a decreased (P=0.30) per cycle 14-d viable pregnancy rate (14 31 , 45.2%) compared with that of endophyte-free grazing (12 16 , 75.0%). Early embryonic death rates were higher (P=0.20) in the endophyte-infected group (6 20 , 30.0%) than the endophyte-free group (1 13 , 7.7%). Cumulative pregnancy rates after a 60-d breeding period did not differ between the 2 groups. Embryonic development based on mean vesicle height at 14-d was not significantly different between treatment groups for embryos that maintained viability. Embryos that underwent early embryonic death were smaller (P<0.10) at Day-14 than embryos that maintained viability. Mean plasma progesterone concentrations were significantly (P< 0.01) greater at Day-21 postovulation in endophyte-infected mares in which the embryo remained viable (15.8 ng/ml) than in endophyte-free mares that experienced early embryonic death (9.8 ng/ml) or that demonstrated prolongation of luteal function (11.2 ng/ml). The results of this study suggest that grazing endophyte-infected tall fescue can have a detrimental effect on reproductive efficiency in the mare due to an increase in cycles bred per pregnancy rate, increased early embryonic death rate and prolongation of luteal function.