STATISTICAL ASPECTS OF THE INDEPENDENT JOINT ACTION OF POISONS, PARTICULARLY INSECTICIDES

Methods are described for discovering whether a mixture of two poisons is as toxic as predicted on the hypothesis of independent joint action. These include X²tests and a procedure for finding the maximum likelihood estimate of the coefficient of correlation in resistance to the poisons. The methods are illustrated using data from insecticidal tests.
In the insecticidal tests, flour beetles, Tribolium castaneum Herbst, were sprayed with, or exposed to films of, different insecticides in solution in Shell oil P 31. The insecticides were pyrethrins, D.D.T., and B.H.C., singly and in pairs. The statistical analysis of the results showed that pyrethrins and D.D.T. could have acted independently both as films and as direct sprays; D.D.T. and B.H.C. could have acted independently as films but not as direct sprays; and B.H.C. and pyrethrins could not have acted independently either as films or as direct sprays.
These findings are discussed, and it is concluded that independent action should be regarded as a special case of a more general type of joint action for which the term dissimilar is proposed. A general method of approach is suggested for the conception and development of hypotheses of the joint action of poisons.     

THE BIOLOGY OF THE RICE WEEVIL, CALANDRA ORYZAE (L.)

The length of the developmental instars of Calandra oryzae has been estimated at relative humidities of 50, 60, 70 and 80% at 21° C., at 70% at 18, 25 and 30° C., and at 80% at 18° C., by the dissection daily of samples of infested wheat-grains. The results are expressed as a median obtained by the method of probit analysis. Comparison of this method, with estimates of median and mean obtained by orthodox arithmetical methods from similar work on Rhizopertha , show that the probit method gives good estimates.
About 90% of the eggs laid are fertile. Normally only one adult will develop in a grain, all other individuals being destroyed by cannibalism. The sex ratio is unity. It was not possible to cross C. oryzae and C. granaria.
The daily oviposition rate of C. oryzae at 17, 21 and 25° C. increases with relative humidity. There is a critical point at about 60% r.h . below which egg laying declines rapidly, and mortality is high. At 100% r.h . the oviposition rates per female per day are approximately 1.3, 2.5 and 3.4 at 17, 21 and 25° C. respectively.
In experimental conditions most eggs per grain are obtained by giving isolated females one grain each, but more eggs are laid by females given more than one grain. Daily egg output is reduced by grouping females or including males. In culture, depths of grain up to 7 cm. do not discourage egg laying.     

Repellent and insecticidal activities of essential oils from Artemisia princeps and Cinnamomum camphora and their effect on seed germination of wheat and broad bean

Repellent and insecticidal activities of essential oils extracted from leaves of Artemisia princeps Pamp and seeds of Cinnamomum camphora (L.) Presl. against storage pests Sitophillus oryzae L. and Bruchus rugimanus Bohem were investigated. Results showed that the two individual oils displayed good, but their mixture (1:1) exhibited much better repellent activities at concentrations from 250 to 1000 microg g(-1) and insecticidal actions at concentrations 1000 microg g(-1) against the test beetles S. oryzae and B. rugimanus. Oils from A. princeps and C. camphora applied individually were significantly toxic to seed germination of wheat at 500 microg ml(-1). However, no toxic effects were found when the two oils were mixed (1:1 w/w) at the same concentration. These observations indicated that the mixture of the two plant-derived oils had a synergic effect and could be used in the control of storage pests.     

The phytotoxicity of D.D.T. and of benzene hexachloride

Spraying with two preparations of D.D.T. had a negligible effect on the maturation of outdoor tomatoes and of summer cabbage. Spraying with a commercial D.D.T. suspension damaged cucurbits and treatment with the suspension base distorted glasshouse cucumbers during one season only.
Benzene hexachloride (B.H.C.) sprays did not affect the yield or flavour of tomatoes, but, at concentrations higher than those normally used, they severely scorched radish, turnip, swede, kale, spinach and beetroot seedlings. The application of B.H.C. dusts stunted seedlings of radish.
On one occasion young potato foliage was damaged by relatively high concentrations of two compounded B.H.C. sprays and by B.H.C. dust but late-season older foliage was unaffected. A spray compounded from B.H.C. dispersible powder caused tainting of potatoes, peas, carrots, beetroot, marrows, cauliflowers and lettuce. Apples and plums treated with B.H.C. spray preparations developed a taint after cooking.     

THE INSECTICIDAL ACTIVITY OF PARATHION, ITS ISOMERS AND SOME RELATED COMPOUNDS

Heat treatment of parathion at 140° C. and above resulted in isomerization and then thermal decomposition; the loss of toxicity to Calandra granaria being correlated with a reduction of the thiono-sulphur content. Similar results were obtained with O:O-dimethyl-(4-nitrophenyl)-thiophosphate. Paraoxon, O:S-diethyl-(4-nitrophenyl)-thiophosphate, and O:O-diethyl-S-(4-nitrophenyl)-thiophosphate, although all possessing considerable contact activity, were less insecticidal than parathion; O:O-dimethyl-(4-nitrophenyl)-thiophosphate, on the other hand, was considerably more effective than parathion. O:O- bis (2-chloroethyl)-O-(4-nitrophenyl)-thiophosphate was considerably less toxic than parathion and appeared to have a different mode of action. Compounds, S-ethyl- bis -(4-nitrophenyl)-thiophosphate, O-ethyl- bis -(4-nitrophenyl)-thiophosphate, and triethyl thiophosphate were ineffective as contact insecticides.
In the series of compounds examined, replacement of the group P = S by P = O or alteration in size of the groups attached to the central phosphorus atom caused a reduction of insecticidal activity.     

Dosage reduction to improve the selectivity of deltamethrin between aphids and coccinellids in cereals

The toxicity of the pyrethroid insecticide deltamethrin to a cereal aphid and a coccinellid beetle predator was assessed. Deltamethrin gave effective aphid control in winter wheat at dose-rates of 6.25, 3.13 and 1.56g a.i./ha. The direct exposure of adultCoccinella septempunctata L. (Coleoptera: Coccinellidae) to spray drops was estimated at a range of positions in a cereal crop canopy from volumetric analysis of fluorescent tracer deposits. These measurements were used to calculate exposure to deltamethrin at the three experimental dose-rates. Observations of coccinellid beetle distribution through a cereal crop canopy permitted a realistic range of direct contact doses tobe calculated and the toxic effects of these levels of exposure to be predicted from laboratory dose-response data. Estimated beetle mortalities from direct exposure were 19, 8 and 3% at the three experimental dose-rates.In situbioassays with adultC. septempunctata which exposed beetles continuously to deltamethrin residues on flag leaves, resulted in 100, 94 and 39% mortality respectively at these dose-rates during the 10 days after spray application. Additionalin situ bioassays exposed beetles to deltamethrin residues on flag leaves for 24 h and then transferred surviving beetles to the soil under the cereal crop canopy for a further 9 days. This resulted in 89, 69 and 29% beetle mortality respectively at the three dose-rates. Mortality predictions combining both direct contact and residual exposure were made for the three dose-rates.to determine the maximum impact of summer sprays of deltamethrin on adult coccinellid populations in cereals. These worst case predictions suggested that a reduction in dose-rate by as much as three quarters of the recommended application rate in UK cereals may be necessary to preserve approximately 60% of adultC. septempunctata in the crop over the 10 days after a deltamethrin spray application. The methodology described may be appropriate for estimating selective dose-rates for key enemies in a range of crops.     

Impact on soil-dwelling arthropods in citrus orchards of spraying horticultural mineral oil, carbaryl or methidathion

Abstract  Impacts of an n C24 horticultural mineral oil (HMO) and two synthetic insecticides (carbaryl and methidathion) on the abundance and species diversity of soil-dwelling arthropods were evaluated in two citrus orchards in the coastal region of New South Wales, Australia. In the first orchard, mature Valencia orange trees were sprayed in summer with one of HMO, carbaryl or methidathion delivered at low (2000 L/ha) or high (10 000 L/ha) volumes; the sprays were applied either once (February) or twice (December and February). HMO had no significant impact but the synthetic insecticides, irrespective of spray frequency or spray volume, significantly reduced the abundance and species diversity of the arthropods including springtails and generalist predators, such as spiders, staphylinid beetles and mesostigmatid mites. In the second orchard, blocks of Washington navel and Valencia orange trees were sprayed with either HMO or methidathion; sprays were applied twice, 2 weeks apart, in late summer (February), first at a rate of 6500 L/ha and second at 5500 L/ha. Results were similar to those in the first orchard, except that the effect of methidathion on carabid beetles and ants was not significant.     

Comparison of bacteriostatic and bactericidal activity of 13 essential oils against strains with varying sensitivity to antibiotics

Aims:  To compare the bacteriostatic and bactericidal activity of 13 chemotyped essential oils (EO) on 65 bacteria with varying sensitivity to antibiotics.
Methods and Results:  Fifty-five bacterial strains were tested with two methods used for evaluation of antimicrobial activity (CLSI recommendations): the agar dilution method and the time-killing curve method. EO containing aldehydes ( Cinnamomum verum bark and Cymbopogon citratus ), phenols ( Origanum compactum , Trachyspermum ammi , Thymus satureioides , Eugenia caryophyllus and Cinnamomum verum leaf) showed the highest antimicrobial activity with minimum inhibitory concentration (MIC) <2% (v/v) against all strains except Pseudomonas aeruginosa . Alcohol-based EO ( Melaleuca alternifolia , Cymbopogon martinii and Lavandula angustifolia ) exhibited varying degrees of activity depending on Gram status. EO containing 1·8-cineole and hydrocarbons ( Eucalyptus globulus , Melaleuca cajeputii and Citrus sinensis ) had MIC_90% ≥ 10% (v/v). Against P. aeruginosa , only C. verum bark and O. compactum presented MIC ≤2% (v/v). Cinnamomum verum bark, O. compactum , T. satureioides , C. verum leaf and M. alternifolia were bactericidal against Staphylococcus aureus and Escherichia coli at concentrations ranging from to 0·31% to 10% (v/v) after 1 h of contact. Cinnamomum verum bark and O. compactum were bactericidal against P. aeruginosa within 5 min at concentrations <2% (v/v).
Conclusions:  Cinnamomum verum bark had the highest antimicrobial activity, particularly against resistant strains.
Significance and Impact of the Study:  Bacteriostatic and bactericidal activity of EO on nosocomial antibiotic-resistant strains.     

Environmental interactions with the toxicity of plant essential oils to the poultry red mite Dermanyssus gallinae

The toxicity of a range of plant essential oils to the poultry red mite, Dermanyssus gallinae (De Geer) (Acari: Dermanyssidae), a serious ectoparasitic pest of laying hens throughout Europe and elsewhere, was assessed in the laboratory. Dermanyssus gallinae may cause losses in egg production, anaemia and, in extreme cases, death of hens. With changes in legislation and consumer demand, alternatives to synthetic acaricides are needed to manage this pest. Fifty plant essential oils were selected for their toxicity to arthropods reported in the literature. Twenty-four of these essential oils were found to kill > 75% of adult D. gallinae in contact toxicity tests over a 24-h period at a rate of 0.21 mg/cm². Subsequent testing at lower rates showed that the essential oils of cade, manuka and thyme were especially toxic to adult D. gallinae . The toxicity of the seven most acaricidal essential oils was found to be stable at different temperatures likely to be encountered in commercial poultry housing (15°C, 22°C and 29°C), although results suggest that humidity and dust might influence the toxicity of some of the oils tested. The toxicity of clove bud essential oil to D. gallinae , for example, was increased at high humidity and dust levels compared with ambient levels. The results suggest that certain essential oils may make effective botanical pesticides for use against D. gallinae , although it is likely that issues relating to the consistency of the toxic effect of some oils will determine which oils will be most effective in practice.     

The Effect of Temperature, Pressure and Chlorine Concentration of Spray Washing Water on Numbers of Bacteria on Lamb Carcases

Combined analysis of three experiments showed that when lamb carcases with initial bacterial numbers of between logi₁₀3.29 and 4.22/cm² were spray washed, statistically significant reductions in bacterial numbers of log₁₀O.5 were obtained when the spray wash water temperature was > 57°C, and reductions of log₁₀1.0 were obtained when the temperature was ≥ 80°C. Reductions at all temperatures were enhanced by log₁₀0.66 when the water contained 30 µg/ml chlorine, but increasing the concentration to 450 µg/ml reduced bacterial numbers only by a further log₁₀0–29. At highly contaminated sites increasing the duration of spraying from 30 to 120 s significantly increased the reductions obtained when water containing added chlorine was used. Reductions in bacterial numbers after spray washing with pressures of 3.5, 5.6. 7.7 kg/cm² were not significantly different.     

Assessing the effects of application time and temperature on the efficacy of hot-water sprays to mitigate fouling by Dreissena polymorpha (zebra mussels Pallas)

Dreissenid mussel (Dreissena polymorpha, Dreissena rostriformis bugensis) expansion into the Western US has renewed interest in hot-water spray mitigation of mussel fouling on boat hulls, trailers, and other equipment. However, the efficacy of hot-water sprays to mitigate dreissenid fouling has not been experimentally assessed. Emersed, adult D. polymorpha were exposed to low-pressure, hot-water sprays at 40, 50, 60, 70, and 80°C for 1, 5, or 10 s. Sprays at ≥60°C for 10 s or 80°C at ≥5 s were 100% lethal. In contrast, 1–10 s exposures did not induce 100% mortality at ≤50°C. The results indicate that mitigation of D. polymorpha fouling, especially in areas protected from the hydraulic impacts of high-pressure sprays requires spray temperatures of > 80°C applied for >5 s or no less than 60°C applied for >10 s. Thus, presently recommended spray temperatures of ≥60°C may not be 100% effective unless applied for >10 s.     

Green peach aphid [Myzus persicae (Sulzer) (Hemiptera: Aphididae)] control using Brassicaceae ethyl ester oil sprays

Control of green peach aphid (Myzus persicae), a globally important pest, using plant‐derived oils is a promising alternative to conventional insecticides. Although various plant‐derived oils are potentially useful for insect control, dose–response studies and efficacy comparisons among oils have not been widely reported. Our objective was to compare M. persicae control by plant‐derived oils, focusing on oils derived from Brassicaceae species that exhibit rotational and environmental quality benefits. We thus applied sprays of emulsified ethyl esters from the seed oils of yellow mustard (Sinapis alba), oriental mustard (Brassica juncea) and rapeseed (Brassica napus) to M. persicae in a laboratory bioassay. A dose–response relationship was modelled for the S. alba spray yielding LD₅₀/LD₉₅ values of 18.2 ± 0.87/128.1 ± 5.10 μg ester per cm² (P < 0.0001). Ethyl esters of oils from all three species and soybean (Glycine max) ethyl ester were compared to determine the efficacy of Brassicaceae oils relative to the dominant plant‐oil spray currently available. All ethyl esters were equally efficacious despite measured differences in fatty acid profiles among the oils. Oils derived from mustards B. juncea and S. alba are potentially useful feedstocks for the production of insecticidal sprays, and testing on additional insects is warranted.     

A note on the effect of sporulation conditions on the resistance of Bacillus spores to heat and chemicals

Spores of Bacillus subtilis SA22 harvested after 22 d incubation on nutrient agar at 30°C were more resistant to 0–04% peracetic acid at 20°C than spores harvested following 2 d incubation. Similarly, spores of B. subtilis globigii B17, harvested after 7 d incubation on a sporulation agar were up to 10 times less resistant to 0.04% peracetic acid at 20°C than spores harvested after 35 d incubation. An increase in resistance to heating at 100°C and to exposure to 17.7% hydrogen peroxide at 20°C occurred as the age of B. subtilis SA22 spores prior to harvesting increased, whereas differences in resistance were not observed with spores of B. subtilis globigii B17.     

Effects of a 4 °C increase in temperature on partitioning of leaf area and dry mass, root respiration and carbohydrates

1. Plants of Bellis perennis, Dactylis glomerata and Poa annua were grown from seed in controlled-environment cabinets at either 16 or 20 °C; at the higher temperature all three species had increased total dry mass and leaf area when assessed on the basis of chronological time. On the basis of thermal time (summation of degree-days above 0 °C; days °C) temperature decreased the dry mass in P. annua.
2. Partitioning was assessed as a change in the allometric coefficients relating shoot and root dry mass, leaf and plant mass, leaf area and plant mass, and leaf area and leaf mass. Of the 12 relationships examined only three were affected by temperature: there was increased partitioning towards the shoot relative to the root in D. glomerata and increased partitioning towards leaf area rather than leaf mass in D. glomerata and B.perennis .
3. Root respiration was unaffected by temperature of growth in D. glomerata and P.annua but was lower in B. perennis grown at elevated temperature.
4. Root respiration acclimated to temperature in P. annua and B. perennis (i.e. when measured at the same temperature, respiration was higher in plants grown at 16 °C).
5. Root soluble carbohydrate concentration was unaffected by temperature of growth in any of the species. Feeding sucrose to the roots for a short period had no effect on the rate of respiration of B. perennis or D. glomerata but increased root respiration of P. annua .     

Thermal inactivation kinetics of Bacillus subtilis spores suspended in buffer and in oils

The heat resistance of Bacillus subtilis 5230 and A spores freeze dried and suspended in buffer or oils was investigated. As expected, spores were more resistant to heat when suspended in oils than in buffer. This was ascribed to the low a _w of oils and to their content of free fatty acids. Linear survivor curves were obtained for spores suspended in buffer at 105°C or above and for B. subtilis A spores suspended in a vegetable oil. However, the survivor curves of the spores suspended in mineral oil (strain 5230) or olive oil (both strains) were concave upward with a characteristic tailing. The tailing could not be ascribed to spore clumping or to a specific heat injury that can be circumvented by Ca-dipicolinate. It is possibly due to another mechanism of injury or to the activation at high temperature of a normally dormant germination system.     

Antimicrobial properties of plant essential oils and essences against five important food-borne pathogens

The antimicrobial properties of 21 plant essential oils and two essences were investigated against five important food-borne pathogens, Campylobacter jejuni, Salmonella enteritidis, Escherichia coli, Staphylococcus aureus and Listeria monocytogenes. The oils of bay, cinnamon, clove and thyme were the most inhibitory, each having a bacteriostatic concentration of 0·075% or less against all five pathogens. In general, Gram-positive bacteria were more sensitive to inhibition by plant essential oils than the Gram-negative bacteria. Campylobacter jejuni was the most resistant of the bacteria investigated to plant essential oils, with only the oils of bay and thyme having a bacteriocidal concentration of less than 1%. At 35 °C, L. monocytogenes was extremely sensitive to the oil of nutmeg. A concentration of less than 0·01% was bacteriostatic and 0·05% was bacteriocidal, but when the temperature was reduced to 4 °C, the bacteriostatic concentration was increased to 0·5% and the bacteriocidal concentration to greater than 1%.     

Induction of embryogenic pollen grains in situ and subsequent in vitro pollen embryogenesis in Nicotiana tabacum by treatments of the pollen donor plants with feminizing agents

Tobacco plants ( Nicotiana tabacum L., var. Badischer Burley) were treated with chemicals (sprays and soil drenches) known to affect sex expression in other species. Their effect was tested on sex balance, pollen sterility, embryogenic pollen grain (P-grain) formation in situ, and on pollen plant formation in anther and pollen cultures after anther preculture. Napthalene acetic acid (NAA) increased the length of pistils and stamens and shifted sex balance towards femaleness when the plants were raised in long or short days at 24 or 15°C. In parallel, pollen sterility, P-grain frequency in situ and pollen plant production from anther and pollen cultures were increased by NAA. Alar 85 redueed the length of pistils and stamens and shifted sex balance towards femaleness when the plants were raised in long days at 24°C, but shifted it towards maleness in short days and/or at 15°C. In parallel, pollen sterility, P-grain frequency in situ, and pollen plant production in vitro were increased when plants in long days at 24°C were treated with Alar 85, but decreased when plants in short days and/or at 15°C were treated. Ethrel, Cycocel, and GA₃ applied in a similar manner, were ineffective. Water sprays and nitrogen starvation shifted sex balance towards femaleness in long days at 15°C and increased pollen sterility, P-grain frequency in situ and pollen plant production in vitro. At 24°C, water sprays and nitrogen starvation had no effect.     

The toxicity of certain aliphatic chlorinated hydrocarbons to Calandra granaria L. and other insects infesting grain

An account is given of the biological data obtained during an investigation into the possibility of using certain aliphatic chlorinated hydrocarbons for fumigation of empty grain sacks infested by insects.
The relative toxicity of three mixtures of chlorinated hydrocarbons to adult Calandra granaria , was ascertained: the best contained 3 parts ethylene dichloride and 1 part tetrachlorethylene by volume. Among seven species of insects likely to occur in empty sacks, the resistance of C. granaria adults to the 3: 1 mixture was exceeded only by that of Trogoderma granarium larvae. The toxicity of the mixture to C. granaria was determined over a range of concentrations and periods of exposure: at 20 C and 70% R.H. a concentration × time product of about 1500 mg.-hr./l. was required for 100% kill. These results were confirmed by inserting tubes containing weevils into bundles of sacks, which were then exposed to fumigant in a steel bin.
The toxicity to adult C. granaria of mixtures containing different proportions of ethylene dichloride and tetrachlorethylene increased as the proportion of ethylene dichloride increased. The toxicity of tetrachlorethylene to the weevils was very low in relation to that of ethylene dichloride.
The concentration × time product for 100 % kill of C. granaria at 20 C. by ethylene dichloride alone was about 1100 mg.-hr./1. The products for 50 % kill of C. granaria at 10, 15, 20 and 25 C were in the proportion of 3.0: 2.0: 1.0: 0.7 respectively.
Weevils which recovered from severe narcosis by the 3: 1 mixture were able to lay viable eggs.     

PETROLEUM OIL FRACTIONS AS PLANT SPRAY MATERIALS THE EFFECT OF REFINING AN OIL UPON ITS DEGREE OF PHYTOTOXICITY

Data are presented showing the effect of degree of refining upon the specific gravity, viscosity, unsulphonatable mineral residue (U.R.), refractive index, average molecular weight, group analysis, sulphur content and phototoxicity of two spindle oil distillates in the boiling range 300–390° C.
Cucumber plants proved very suitable for demonstrating differences in phyto-toxic properties of the oils.
Spindle oils refined to a U.R. of 82 or higher, and a specific gravity of 0.885 or lower, showed no phytotoxicity in the tests on these plants. A fair agreement was found between the results of these tests and the specifications laid down by Dr H. Martin for spray oils.     

Heat resistance of Cronobacter species (Enterobacter sakazakii) in milk and special feeding formula

Aim:  To determine D - and z -values of Cronobacter species ( Enterobacter sakazakii ) in different reconstituted milk and special feeding formula and the effect of reconstitution of powdered milk and special feeding formula with hot water on the survival of the micro-organism.
Methods and Results:  Five Cronobacter species (four C. sakazakii isolates and C. muytjensii ) were heated in reconstituted milk or feeding formula pre-equilibrated at 52–58°C for various times or mixed with powdered milk or feeding formula prior to reconstitution with water at 60–100°C. The D -values of Cronobacter at 52–58°C were significantly higher in whole milk (22·10–0·68 min) than in low fat (15·87–0·62 min) or skim milk (15·30–0·51 min) and significantly higher in lactose-free formula (19·57–0·66 min) than in soy protein formula (17·22–0·63 min). The z -values of Cronobacter in reconstituted milk or feeding formula ranged from 4·01°C to 4·39°C. Water heated to ≥70°C and added to powdered milk and formula resulted in a > 4 log₁₀ reduction of Cronobacter .
Conclusions:  The heat resistance of Cronobacter should not allow the survival of the pathogen during normal pasteurization treatment. The use of hot water (≥70°C) during reconstitution appears to be an effective means to reduce the risk of Cronobacter in these products.
Significance and Impact of the Study:  This study supports existing data available to regulatory agencies and milk producers that recommended heat treatments are sufficient to substantially reduce risk from Cronobacter which may be present in these products.