Hot water immersion for surface disinfestation of Maconellicoccus hirsutus (Homoptera: Pseudococcidae)

Mealybug Maconellicoccus hirsutus (Green) adults, nymphs, crawlers, and eggs were tested for their susceptibility to hot water immersion at 47, 48, and 49 degrees C. Eggs inside ovisacs were found most tolerant with prolonged survival compared with other stages at all temperatures. Ovisacs required an average of 1.38, 1.46, and 1.62 times longer treatment duration than adults, nymphs, and crawlers, respectively, for 99.9% predicted mortality at 47, 48, and 49 degrees C. Lethal time estimations were calculated from inverse predictions of regressions derived from logit-transformed data as well as those created using a kinetic model. LT 99.9 estimations were 47.0, 21.2, and 11.9 min at 47, 48, and 49 degrees C, respectively, by using regressions with logit transformations. The kinetic model predictions were 43.9, 19.6, and 11.1 min at 47, 48, and 49 degrees C, respectively. During the study no emergence from eggs inside ovisacs was found after treatments of 52, 24, and 14 min at 47, 48, and 49 degrees C, respectively. Results from this study provide efficacious temperature-time treatments.     

Life history of the mealybug, Maconellicoccus hirsutus (Hemiptera: Pseudococcidae), at constant temperatures

Important life history parameters of the mealybug, Maconellicoccus hirsutus (Green), were characterized on hibiscus (Hibiscus rosa-sinensis L.) cuttings at six constant temperatures between 15 and 35 degrees C. The development of M. hirsutus was the fastest at 27 degrees C, where the mealybugs completed development in approximately 29 d. The lower (T(min)) and upper (T(max)) developmental thresholds and the optimal developmental temperature (T(opt)) for the development of female mealybugs were estimated as 14.5, 35, and 29 degrees C, respectively. The thermal constant (K), which is the number of temperature-day or degree-day units required for development, of the females was 347 DD. The original distribution range prediction (based on T(min) = 17.5 degrees C and K = 300 DD) indicated that M. hirsutus could complete at least one generation in all of the continental United States. However, results of this study suggested that the distribution range of M. hirsutus may expand northward because of the lower T(min), and the predicted number of generations in a year may be lower because of the higher K required to complete each generation. The average cumulative survival rate of M. hirsutus at 25 and 27 degrees C was 72%, which was significantly higher than 51 and 62% at 20 and 30 degrees C, respectively. M. hirsutus reproduced sexually, with each mated female producing 260-300 eggs between 20 and 27 degrees C but only approximately 100 eggs at 30 degrees C. Female longevity was reduced from 28 d at 20 degrees C to 19-21 d at 25-30 degrees C. At 27 degrees C, the net reproductive rate (R(o)) was estimated at 165 female symbol/female symbol, the intrinsic rate of population increase (r(m)) was 0.119 (female symbol/female symbol/d), the generation time (T(G)) was 43 d, and the doubling time (DT) was 5.8 d. The life table statistics suggested that the currently released biological control agents, which have higher r(m) than M. hirsutus, will be able to complete more generations than the mealybug within the tested temperature range; thus, they are effective against M. hirsutus.     

Susceptibility of Plodia interpunctella (Lepidoptera: Pyralidae) developmental stages to high temperatures used during structural heat treatments

Heating the ambient air of a whole, or a portion of a food-processing facility to 50 to 60 degrees C and maintaining these elevated temperatures for 24 to 36 h, is an old technology, referred to as heat treatment. There is renewed interest in adopting heat treatments around the world as a viable insect control alternative to fumigation with methyl bromide. There is limited published information on responses of the Indian meal moth, Plodia interpunctella (Hübner), exposed to elevated temperatures typically used during heat treatments. Time-mortality relationships were determined for eggs, fifth-instars (wandering-phase larvae), pupae, and adults of P. interpunctella exposed to five constant temperatures between 44 and 52 degrees C. Mortality of each stage increased with increasing temperature and exposure time. In general, fifth-instars were the most heat-tolerant stage at all temperatures tested. Exposure for a minimum of 34 min at 50 degrees C was required to kill 99% of the fifth-instars. It is proposed that heat treatments aimed at controlling fifth-instars should be able to control all other stages of P. interpunctella.     

Thermal death kinetics of red flour beetle (Coleoptera: Tenebrionidae)

While developing radio frequency heat treatments for dried fruits and nuts, we used a heating block system developed by Washington State University to identify the most heat-tolerant life stage of red flour beetle, Tribolium castaneum (Herbst), and to determine its thermal death kinetics. Using a heating rate of 15 degrees C/min to approximate the rapid heating of radio frequency treatments, the relative heat tolerance of red flour beetle stages was found to be older larvae > pupae and adults > eggs and younger larvae. Lethal exposure times for temperatures of 48, 50, and 52 degrees C for the most heat-tolerant larval stage were estimated using a 0.5th order kinetic model. Exposures needed for 95% mortality at 48 degrees C were too long to be practical (67 min), but increasing treatment temperatures to 50 and 52 degrees C resulted in more useful exposure times of 8 and 1.3 min, respectively. Red flour beetle was more sensitive to changes in treatment temperature than previously studied moth species, resulting in red flour beetle being the most heat-tolerant species at 48 degrees C, but navel orangeworm, Amyelois transitella (Walker), being most heat tolerant at 50 and 52 degrees C. Consequently, efficacious treatments for navel orangeworm at 50-52 degrees C also would control red flour beetle.     

Time-mortality relationships for Tribolium castaneum (Coleoptera: Tenebrionidae) life stages exposed to elevated temperatures

The use of elevated temperatures (> or = 40-60 degrees C) or heat treatments for managing insects in food-processing facilities is a viable alternative to space fumigation with methyl bromide. Quantitative data are lacking on the responses of life stages of the red flour beetle, Tribolium castaneum (Herbst), an important pest of food-processing facilities worldwide, to elevated temperatures used during heat treatments. We determined time-mortality relationships for eggs, young (neonate) larvae, old larvae, pupae, and adults of T. castaneum, exposed to constant temperatures of 42, 46, 50, 54, 58, and 60 degrees C. Generally, mortality of each stage increased with an increase in temperature and exposure time. Young larvae were the most heat-tolerant stage, especially at temperatures > or = 50 degrees C. Exposure for a minimum of 7.2 h at > or = 50 degrees C was required to kill 99% of young larvae, whereas the other stages required < or = 1.8 h. Heat treatments that control young larvae should control all other stages of T. castaneum, and young larvae should be used as test insects to evaluate efficacy against T. castaneum during an actual facility heat treatment. These results provide the basis for successful use of elevated temperatures for management of T. castaneum life stages associated with food-processing facilities.     

Susceptibility of Maconellicoccus hirsutus (Homoptera: Pseudococcidae) to methyl bromide

Eggs, crawlers, early nymphs, late nymphs, and adults of the pink hibiscus mealybug, Maconellicoccus hirsutus (Green), were tested for their susceptibility to methyl bromide in 2-h laboratory fumigations at ambient conditions (25 degrees C, 95% RH). Dose-response tests indicated that the egg was the most susceptible life stage with an LC99 of 20.2 mg/liter. Based on probit analysis of dose-response data, no significant differences were observed among susceptibilities of the crawler, early stage or late stage nymphs, or adults at either the LC50 or LC99 level, but late stage nymphs were more tolerant than early stage nymphs in a separate paired comparison test. Confirmatory tests showed that a dose of 48 mg/liter methyl bromide, the USDA-Animal Plant Health Inspection Service treatment dose schedule for mealybugs at 21-26 degrees C, produced 100% mortality of all life stages. On the basis of these results, we conclude that the methyl bromide treatment schedule for mealybugs will provide quarantine security for M. hirsutus infesting commodities for export or import.     

Thermotolerance induced by heat, sodium arsenite, or puromycin: its inhibition and differences between 43 degrees C and 45 degrees C

When CHO cells were treated either for 10 min at 45-45.5 degrees C or for 1 hr with 100 microM sodium arsenite (ARS) or for 2 hr with 20 micrograms/ml puromycin (PUR-20), they became thermotolerant to a heat treatment at 45-45.5 degrees C administered 4-14 hr later, with thermotolerance ratios at 10(-3) isosurvival of 4-6, 2-3.2, and 1.7, respectively. These treatments caused an increase in synthesis of HSP families (70, 87, and 110 kDa) relative to total protein synthesis. However, for a given amount of thermotolerance, the ARS and PUR-20 treatments induced 4 times more synthesis than the heat treatment. This decreased effectiveness of the ARS treatment may occur because ARS has been reported to stimulate minimal redistribution of HSP-70 to the nucleus and nucleolus. Inhibiting protein synthesis with cycloheximide (CHM, 10 micrograms/ml) or PUR (100 micrograms/ml) after the initial treatments greatly inhibited thermotolerance to 45-45.5 degrees C in all cases. However, for a challenge at 43 degrees C, thermotolerance was inhibited only for the ARS and PUR-20 treatments. CHM did not suppress heat-induced thermotolerance to 43 degrees C, which was the same as heat protection observed when CHM was added before and during heating at 43 degrees C without the initial heat treatment. These differences between the initial treatments and between 43 and 45 degrees C may possibly be explained by reports that show that heat causes more redistribution of HSP-70 to the nucleus and nucleolus than ARS and that redistribution of HSP-70 can occur during heating at 42 degrees C with or without the presence of CHM. Heating cells at 43 degrees C for 5 hr after thermotolerance had developed induced additional thermotolerance, as measured with a challenge at 45 degrees C immediately after heating at 43 degrees C. Compared to the nonthermotolerant cells, thermotolerance ratios were 10 for the ARS treatment and 8.5 for the initial heat treatment. Adding CHM (10 micrograms/ml) or PUR (100 micrograms/ml) to inhibit protein synthesis during heating at 43 degrees C did not greatly reduce this additional thermotolerance. If, however, protein synthesis was inhibited between the initial heat treatment and heating at 43 degrees C, protein synthesis was required during 43 degrees C for the development of additional thermotolerance to 45 degrees C.(ABSTRACT TRUNCATED AT 400 WORDS)     

Thermal inactivation of Enterococcus faecium: effect of growth temperature and physiological state of microbial cells

AIMS: To provide data on the effects on culture temperature and physiological state of cells on heat resistance of Enterococcus faecium, which may be useful in establishing pasteurization procedures. METHODS AND RESULTS: The heat resistance of this Ent. faecium (ATCC 49624 strain) grown at different temperatures was monitored at various stages of growth. In all cases, the bacterial cells in the logarithmic phase of growth were more heat sensitive. For cells which had entered in the stationary phase, D70 values of 0.53 min at 5 degrees C, 0.74 min at 10 degrees C, 0.83 min at 20 degrees C, 0.79 min at 30 degrees C, 0.63 min at 37 degrees C, 0.48 min at 40 degrees C and 0.41 min at 45 degrees C were found. By extending the incubation times cells were more heat resistant as stationary phase progressed, although a different pattern was observed for cells grown at different temperatures. At the lower temperatures heat resistance increased progressively, reaching D70 values of 1.73 min for cells incubated at 5 degrees C for 50 days and 1.04 min for those grown at 10 degrees C for 16 days. At other temperatures assayed heat resistance became stable for late stationary phase cells, reaching D70 values of 1.05, 1.08 and 1.01 min for cultures incubated at 20, 30 and 37 degrees C. Heat resistance of cells obtained at higher temperatures, 40 and 45 degrees C, was significantly lower, with D70 values of 0.76 and 0.67 min, respectively. Neither the growth temperature nor the growth phase modified the z-values significantly. CONCLUSIONS: D70 values obtained for Ent. faecium (ATCC 49624) varies from 0.33 to 1.73 min as a function of culture temperature and physiological state of cells. However, z values calculated were not significantly influenced by these factors. A mean value of 4.50 +/- 0.39 degrees C was found. SIGNIFICANCE AND IMPACT OF THE STUDY: Overall results strongly suggest that, to establish heat processing conditions of pasteurized foods ensuring elimination of Ent. faecium, it is advisable to take into account the complex interaction of growth temperature and growth phase of cells acting on bacterial thermal resistance.     

Thermal treatments to increase acoustic detectability of Sitophilus oryzae (Coleoptera: Curculionidae) in stored grain.

Hidden infestations of stored-product insect larvae are detected most rapidly by acoustic techniques when the larvae are highly active. Larval activity is periodic, however, and it tends to decrease after the larvae are disturbed or cooled. Because of the practical need for rapid inspection of grain at commercial elevators, several heat treatments were tested as potential methods of increasing larval activity and improving the speed and reliability of acoustic detection under adverse conditions. Samples of grain infested with 4th instars of Sitophilus oryzae (L.) were exposed to different radiant and convective heat treatments after they had been conditioned at 11 degrees C, 17 degrees C, or room temperature for 12-24 h. Relative activity levels were evaluated over periods of 0-12 h based on the mean levels in a 15-min interval, 2 h after the beginning of a trial. In comparisons among treatments with precooled larvae, relative activity levels 5-10 min after brief heat pulses were 2-30 times higher than activity levels in precooled controls exposed only to ambient temperatures (25 degrees C). After 15-25 min, the relative activity levels of these heated larvae remained 2-5 times higher than those of the ambient controls. Brief movement disturbances inhibited activity for approximately 20 min at any temperature. These results suggest that, in general, larval detectability is enhanced if cool grain samples are warmed and all samples are left undisturbed for 15-20 min before inspection.     

Influence of hyperthermia on gamma-ray-induced mutation in V79 cells

Asynchronously growing V79 cells were assayed for mutation induction following exposure to hyperthermia either immediately before or after being irradiated with 60Co gamma rays. Hyperthermia exposures consisted of either 43.5 degrees C for 30 min or 45 degrees C for 10 min. Each of these heat treatments resulted in a survival level of 42%. For all sequences of combined treatment with hyperthermia and radiation, cell killing by gamma rays was enhanced. Mutation induction by gamma rays was enhanced when heat preceded gamma irradiation, but no increase was observed when heat was given after gamma exposures. Treatment at 45 degrees C for 10 min gave a higher yield in mutants at all gamma doses studied compared to treatment at 43.5 degrees C for 30 min. When heat-treated cells were incubated for different periods before being exposed to gamma rays, thermal enhancement of radiation killing was lost after 24 h. In contrast, only 5-6 h incubation was needed for loss of mutation induction enhancement.     

Susceptibility of Lasioderma serricorne (Coleoptera: Anobiidae) life stages to elevated temperatures used during structural heat treatments

Heat treatment of food-processing facilities involves using elevated temperatures (50-60 degrees C for 24-36 h) for management of stored-product insects. Heat treatment is a viable alternative to the fumigant methyl bromide, which is phased out in the United States as of 2005 because of its adverse effects on the stratospheric ozone. Very little is known about responses of the cigarette beetle, Lasioderma serricorne (F.) (Coleoptera: Anobiidae), a pest associated with food-processing facilities, to elevated temperatures. Responses of L. serricorne life stages to elevated temperatures were evaluated to identify the most heat-tolerant stage. Exposure of eggs, young larvae, old larvae, and adults during heat treatment of a food-processing facility did not clearly show a life stage to be heat tolerant. In the laboratory, exposure of eggs, young larvae, old larvae, pupae, and adults at fixed times to 46, 50, and 54 degrees C and 22% RH indicated eggs to be the most heat-tolerant stage. Time-mortality responses at each of these three temperatures showed that the time for 99% mortality (LT99) based on egg hatchability and egg-to-adult emergence was not significantly different from one another at each temperature. Egg hatchability alone can be used to determine susceptibility to elevated temperatures between 46 and 54 degrees C. The LT99 based on egg hatchability and egg-to-adult emergence at 46 degrees C was 605 and 598 min, respectively, and it decreased to 190 and 166 min at 50 degrees C and 39 and 38 min at 54 degrees C. An exponential decay equation best described LT99 as a function of temperature for pooled data based on egg hatchability and egg-to-adult emergence. Our results suggest that during structural heat treatments eggs should be used in bioassays for gauging heat treatment effectiveness, because treatments aimed at controlling eggs should be able to control all other L. serricorne life stages.     

Thermal responses of oriental fruit fly (Diptera: Tephritidae) late third instars: mortality, puparial morphology, and adult emergence

Responses of late third instars of the oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), to high temperatures (43, 46, and 48 degrees C) were investigated. The different heat exposures not only affected the timing of death but also induced different quantities of malformed puparia and changed the average eclosion time. A majority of larvae died immediately (as larvae) after 30 min at 46 degrees C and > or =15 min at 48 degrees C, whereas most individuals died as pupae after 10-25 min of 46 degrees C, 5-10 min of 48 degrees C, and 40-60 min of 43 degrees C treatments. Lethal times estimated by immediate mortality were longer than those estimated by delayed mortality at the same high temperature. Surviving larvae formed four types of puparial morphology (normal, bottlenose, larviform, and peanut form). The percentage of normal puparia showed a negative correlation with exposure time at all test temperatures. The number of bottlenose was more than the larviform and the peanut at 46 degrees C for < or =20 min and at 48 degrees C for < or =10 min, respectively, whereas the number of larviform was more than the bottlenose and the peanut at 46 degrees C and 48 degrees C for longer exposure times. The average eclosion time increased at first, then decreased as the exposure time prolonged, and the longest average eclosion time occurred in the 40-min exposure at 43 degrees C, 15-min exposure at 46 degrees C, and 10-min exposure at 48 degrees C.     

Assessment of the cell viability of cultured Perkinsus marinus (Perkinsea), a parasitic protozoan of the Eastern oyster, Crassostrea virginica, using SYBRgreen-propidium iodide double staining and flow cytometry

A flow cytometry (FCM) assay using SYBRgreen and propidium iodide double staining was tested to assess viability and morphological parameters of Perkinsus marinus under different cold- and heat-shock treatments and at different growth phases. P. marinus meront cells, cultivated at 28 degrees C, were incubated in triplicate for 30 min at -80 degrees C, -20 degrees C, 5 degrees C, and 20 degrees C for cold-shock treatments and at 32 degrees C, 36 degrees C, 40 degrees C, 44 degrees C, 48 degrees C, 52 degrees C, and 60 degrees C for heat-shock treatments. A slight and significant decrease in percentage of viable cells (PVC), from 93.6% to 92.7%, was observed at -20 degrees C and the lowest PVC was obtained at -80 degrees C (54.0%). After 30 min of heat shocks at 40 degrees C and 44 degrees C, PVC decreased slightly but significantly compared to cells maintained at 28 degrees C. When cells were heat shocked at 48 degrees C, 52 degrees C, and 60 degrees C heavy mortality occurred and PVC decreased to 33.8%, 8.0%, and 3.4%, respectively. No change in cell complexity and size was noted until cells were heat shocked at >or=44 degrees C. High cell mortality was detected at stationary phase of P. marinus cell culture. Cell viability dropped below 40% in 28-day-old cultures and ranged 11-25% in 38 to 47-day-old cultures. Results suggest that FCM could be a useful tool for determining viability of cultured P. marinus cells.     

The effect of quick-freezing in ethylene glycol on morphological survival and in vitro development of mouse embryos frozen at different preimplantation stages

This study was conducted to examine the effect of a quick-freezing protocol on morphological survival and in vitro development of mouse embryos cryopreserved in ethylene glycol (EG) at different preimplantation stages. One-cell embryos were harvested from 6-to 8-wk-old CB6F1 superovulated mice, 20 to 23 h after pairing with males of the same strain and hCG injection. The embryos were cultured in human tubal fluid (HTF) containing 4 mg/ml BSA under mineral oil at 37 degrees C in 5% CO(2) plus 95% room air at maximal humidity. Twenty-four to 96 h after collection, the embryos were removed from culture and frozen at the 2 cell, 4 to 8-cell, compact morula, early blastocyst, expanding blastocyst and expanded blastocyst stages. To perform the quick-freeze procedure, embryos were equilibrated in Dulbecco's phosphate buffered saline (DPBS) + 10 % fetal bovine serum (FBS) + 0.25 M sucrose + 3 M ethylene glycol (freeze medium) for 20 min at room temperature (22 to 26 degrees C) and loaded in a single column of freeze medium into 0.25-ml straws (4 to 5 embryos per straw). The straws were held in liquid nitrogen vapor for 2 min and immersed in liquid nitrogen. Embryos were thawed by gentle agitation in a 37 degrees C water bath for 20 sec and transferred to DPBS + 10 % FBS + 0.5 M sucrose (re-hydration medium) for 10 min at room temperature, rinsed 2 times in HTF plus 4 mg/ml BSA and then cultured for 24 to 96 h. Survival of embryos was based on their general morphological appearance after thawing and their ability to continue development upon subsequent culture in vitro. Survival of blastocysts after thawing also required expansion or reexpansion of the blastocoel after several hours in culture. Significant differences were found in the survival and development of mouse embryos at different developmental stages quick-frozen in ethylene glycol and sucrose: 2-cell embryos 43/84 (51%), 4 to 8-cell embryos 44/94 (47%), morulae and early blastocysts 56/70 (80%; P相似文献   

Relative susceptibility of Tribolium confusum life stages exposed to elevated temperatures

Methyl bromide, a space fumigant used in food-processing facilities, may be phased out in the United States by 2005. The use of elevated temperatures or heat treatment is gaining popularity as a methyl bromide alternative. During heat treatment, the temperature of the whole food-processing facility, or a portion of it, is raised and held between 50 and 60 degrees C for 24-36 h to kill stored-product insects. We determined time-mortality responses of the confused flour beetle, Tribolium confusum (Jacquelin du Val), eggs, young larvae, old larvae, pupae, and adults exposed to six constant temperatures between 46 and 60 degrees C. Responses of all five insect stages also were measured using exposure times of 160, 40, and 12 min at 46, 50, and 60 degrees C, respectively. Time-mortality responses of all T. confusum life stages increased with an increase in exposure time and temperature. Both time-mortality and fixed time responses showed eggs and young larvae to be most susceptible at elevated temperatures and old larvae to be least susceptible. Our results suggest that old larvae should be used as test insects to gauge heat treatment effectiveness, because heat treatment aimed at controlling old larvae should be able to control all other T. confusum life stages. Besides providing baseline data for successful use of heat treatments, time-mortality data collected at the six temperatures can be used for developing thermal death kinetic models for this species to predict mortality during actual facility heat treatments.     

Inactivation of Mycobacterium paratuberculosis in cows' milk at pasteurization temperatures.

The thermal inactivation of 11 strains of Mycobacterium paratuberculosis at pasteurization temperatures was investigated. Cows' milk inoculated with M. paratuberculosis at two levels (10(7) and 10(4) CFU/ml) was pasteurized in the laboratory by (i) a standard holder method (63.5 degrees C for 30 min) and (ii) a high-temperature, short-time (HTST) method (71.7 degrees C for 15 s). Additional heating times of 5, 10, 15, 20, and 40 min at 63.5 degrees C were included to enable the construction of a thermal death curve for the organism. Viability after pasteurization was assessed by culture on Herrold's egg yolk medium containing mycobactin J (HEYM) and in BACTEC Middlebrook 12B radiometric medium supplemented with mycobactin J and sterile egg yolk emulsion. Confirmation of acid-fast survivors of pasteurization as viable M. paratuberculosis cells was achieved by subculture on HEYM to indicate viability coupled with PCR using M. paratuberculosis-specific 1S900 primers. When milk was initially inoculated with 10(6) to 10(7) CFU of M. paratuberculosis per ml, M. paratuberculosis cells were isolated from 27 of 28 (96%) and 29 of 34 (85%) pasteurized milk samples heat treated by the holder and HTST methods, respectively. Correspondingly, when 10(3) to 10(4) CFU of M. paratuberculosis per ml of milk were present before heat treatment, M. paratuberculosis cells were isolated from 14 of 28 (50%) and 19 of 33 (58%) pasteurized milk samples heat treated by the holder and HTST methods, respectively. The thermal death curve for M. paratuberculosis was concave in shape, exhibiting a rapid initial death rate followed by significant "tailing." Results indicate that when large numbers of M. paratuberculosis cells are present in milk, the organism may not be completely inactivated by heat treatments simulating holder and HTST pasteurization under laboratory conditions.     

Hot-water treatments for control of Planococcus ficus (Homoptera: Pseudococcidae) on dormant grape cuttings

Hot-water immersions were tested for control of mealybug Planococcus ficus (Signoret), on dormant grape cuttings used for nursery stock. A range of hot-water temperatures (47-58 degrees C) were evaluated at immersion periods of 2, 5, 10, or 20 min, by using a total of 353,720 mealybugs across all treatments. A 5-min immersion at 51 degrees C is effective in killing > 99% of P. ficus. At or above this immersion period and temperature, there was no difference in mealybug stage mortality. We evaluated a commercial operation, which used a 5-min immersion in each of three water tanks: preheating (30.0 +/- 3 degrees C), hot-water (52.8 +/- 0.3 degrees C), and cooling (23 +/- 3 degrees C). The commercial procedure provided 99.8-100% mealybug control in each of three separate trials.     

Effect of commercial-scale high-temperature, short-time pasteurization on the viability of Mycobacterium paratuberculosis in naturally infected cows' milk

Raw cows' milk naturally infected with Mycobacterium paratuberculosis was pasteurized with an APV HXP commercial-scale pasteurizer (capacity 2,000 liters/h) on 12 separate occasions. On each processing occasion, milk was subjected to four different pasteurization treatments, viz., 73 degrees C for 15 s or 25 s with and without prior homogenization (2,500 lb/in(2) in two stages), in an APV Manton Gaulin KF6 homogenizer. Raw and pasteurized milk samples were tested for M. paratuberculosis by immunomagnetic separation (IMS)-PCR (to detect the presence of bacteria) and culture after decontamination with 0.75% (wt/vol) cetylpyridinium chloride for 5 h (to confirm bacterial viability). On 10 of the 12 processing occasions, M. paratuberculosis was detectable by IMS-PCR, culture, or both in either raw or pasteurized milk. Overall, viable M. paratuberculosis was cultured from 4 (6.7%) of 60 raw and 10 (6.9%) of 144 pasteurized milk samples. On one processing day, in particular, M. paratuberculosis appeared to have been present in greater abundance in the source raw milk (evidenced by more culture positives and stronger PCR signals), and on this occasion, surviving M. paratuberculosis bacteria were isolated from milk processed by all four heat treatments, i.e., 73 degrees C for 15 and 25 s with and without prior homogenization. On one other occasion, surviving M. paratuberculosis bacteria were isolated from an unhomogenized milk sample that had been heat treated at 73 degrees C for 25 s. Results suggested that homogenization increases the lethality of subsequent heat treatment to some extent with respect to M. paratuberculosis, but the extended 25-s holding time at 73 degrees C was found to be no more effective at killing M. paratuberculosis than the standard 15-s holding time. This study provides clear evidence that M. paratuberculosis bacteria in naturally infected milk are capable of surviving commercial high-temperature, short-time pasteurization if they are present in raw milk in sufficient numbers.     

Isolation and analysis of thermotolerant mutants of wheat

Thermotolerant mutants of wheat cv. Guardian were isolated by selecting survivors from 5-d-old seedlings of M2 populations exposed to 47 degrees C for 90 min. Progeny testing, using triphenyl tetrazolium chloride reduction as a measure of tissue viability, following heat stress treatment for 120 min at each of three temperatures (32, 38 and 50 degrees C), confirmed the thermotolerant nature of seedlings of 13 mutants. Mutants were isolated at frequencies of 0.1% and 0.2% following the use of sodium azide and ethyl methanesulphonate, respectively. The relative thermotolerance of ten of the mutants and 'Guardian' was then tested by exposing plants to heat stresses of 38 degrees C for 6 h in every 24 h for five successive days at one of four growth stages between seedling and anthesis. Pmax (light-saturated net photosynthetic rate) and chlorophyll content were compared in stressed and unstressed plants. The Pmax of 'Guardian' was depressed by at least 23% by heat stress at each stage; this inhibition was least at ear emergence and greatest at anthesis, the latter being associated with reduced sink size as a result of lowered seed set. The stress-induced inhibition of Pmax in 'Guardian' plants at anthesis had not recovered 3 d after removal of the stress. Mutant lines exhibited different developmental profiles of Pmax thermostability. Mutant tht (thermotolerant) 10, for example, exhibited partial thermostability at each growth stage tested while the Pmax of mutant tht 2 was completely unaffected by heat stress at second node and ear emergence, but was as inhibited as that of 'Guardian' at anthesis; heat stress applied at anthesis in tht 2, but not tht 10, was associated with reduced seed set. Generally, the inhibitory effect of heat stress on Pmax in the mutants was reflected in declines in chlorophyll content. The ten mutants were grouped into nine categories, on the basis of thermotolerance characteristics.