We present a method for calculating the Acute Insecticide Toxicity Loading (AITL) on US agricultural lands and surrounding areas and an assessment of the changes in AITL from 1992 through 2014. The AITL method accounts for the total mass of insecticides used in the US, acute toxicity to insects using honey bee contact and oral LD 50 as reference values for arthropod toxicity, and the environmental persistence of the pesticides. This screening analysis shows that the types of synthetic insecticides applied to agricultural lands have fundamentally shifted over the last two decades from predominantly organophosphorus and N-methyl carbamate pesticides to a mix dominated by neonicotinoids and pyrethroids. The neonicotinoids are generally applied to US agricultural land at lower application rates per acre; however, they are considerably more toxic to insects and generally persist longer in the environment. We found a 48- and 4-fold increase in AITL from 1992 to 2014 for oral and contact toxicity, respectively. Neonicotinoids are primarily responsible for this increase, representing between 61 to nearly 99 percent of the total toxicity loading in 2014. The crops most responsible for the increase in AITL are corn and soybeans, with particularly large increases in relative soybean contributions to AITL between 2010 and 2014. Oral exposures are of potentially greater concern because of the relatively higher toxicity (low LD 50 s) and greater likelihood of exposure from residues in pollen, nectar, guttation water, and other environmental media. Using AITL to assess oral toxicity by class of pesticide, the neonicotinoids accounted for nearly 92 percent of total AITL from 1992 to 2014. Chlorpyrifos, the fifth most widely used insecticide during this time contributed just 1.4 percent of total AITL based on oral LD 50 s. Although we use some simplifying assumptions, our screening analysis demonstrates an increase in pesticide toxicity loading over the past 26 years, which potentially threatens the health of honey bees and other pollinators and may contribute to declines in beneficial insect populations as well as insectivorous birds and other insect consumers.

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Freeze-fracture electron microscopy was used to study further the changes in chlorosome structure during the development of the photosynthetic apparatus in Chloroflexus aurantiacus J-10-fl. During development, in response to decreased light intensity or lower oxygen tension, the number of chlorosomes per cell increased. The same conditions also led to a general thickening of chlorosomes but did not affect their length or width. The thickening of the chlorosomes paralleled increases in the bacteriochlorophyll c/bacteriochlorophyll a ratio. Semiaerobic induction of the photosynthetic apparatus did not produce a synchronous assembly of chlorosomes in all cells of a given culture. Even adjacent cells of a single filament showed great variations in the rate and extent of response. Parallel appearance of (i) approximately 5-nm particles (in a lattice configuration) in the membrane attachment site, (ii) the crystalline baseplate material (with a periodicity of approximately 6 nm) adjacent to the membrane attachment site, and (iii) the chlorosome envelope layer preceded addition of longitudinally oriented, rodlike elements (diameter, congruent to 6 m) to the chlorosome core. It is estimated that each chlorosome can funnel energy into approximately 100 reaction centers. Chlorosomes could be isolated by a simple density gradient procedure only from cells grown at low light intensity. A bacteriochlorophyll a species absorbing at 790 nm was associated with isolated chlorosomes. Lithium dodecyl sulfate-polyacrylamide gel electrophoresis of chlorosomes showed only a few low-molecular-weight polypeptides (less than 15,000).

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The substrate-cytochrome P-450C-21 binding reaction has been investigated in detail by using the purified cytochrome. The apparent substrate dissociation constant (KDapp) depended on the enzyme concentration, indicating that the binding reaction does not follow simple two-component mass action equilibrium. However, the binding data fit reasonably well to a model in which the P-450C-21 exists in a monomer-dimer equilibrium and the substrate does not bind to the dimer. The intrinsic dissociation constant (K1) and the dissociation constant for the dimerization reaction (K2) were calculated from the titration data by a pattern search procedure. K1 and K2 were found to be essentially independent of the enzyme concentration, indicating the appropriateness of the assumed model. In the present study, all factors that increased the dissociation of the dimer, as indicated by an increase in K2, decreased KDapp so that it approached the intrinsic constant K1. These results suggest that there is mutual interaction of the substrate binding and self-association reactions of cytochrome P-450C-21 in the purified preparation.