An extended multiplex PCR method was established to rapidly identify and classify Bacillus thuringiensis strains containing cry (crystal protein) genes toxic to species of Lepidoptera, Coleoptera, and Diptera. The technique enriches current strategies and simplifies the initial stages of large-scale screening of cry genes by pinpointing isolates that contain specific genes or unique combinations of interest with potential insecticidal activities, thus facilitating subsequent toxicity assays. Five pairs of universal primers were designed to probe the highly conserved sequences and classify most (34 of about 60) genes known in the following groups: 20 cry1, 3 cry2, 4 cry3, 2 cry4, 2 cry7, and 3 cry8 genes. The DNA of each positive strain was probed with a set of specific primers designed for 20 of these genes and for cry11A. Twenty-two distinct cry-type profiles were identified from 126 field-collected B. thuringiensis strains. Several of them were found to be different from all published profiles. Some of the field-collected strains, but none of the 16 standard strains, were positive for cry2Ac. Three standard and 38 field-collected strains were positive by universal primers but negative by specific primers for all five known genes of cry7 and cry8. These field-collected strains seem to contain a new gene or genes that seem promising for biological control of insects and management of resistance.

Pollinator decline is acknowledged worldwide and constitutes a major subject of environmental research. Nevertheless, farmers' efforts to protect pollinators in agricultural lands remain very limited, in particular if no compensation scheme is applicable. Current research focuses on measuring pollinator diversity in different landscapes, but research on income gains, due to habitat enhancement and high pollinator diversity, may have greater potential to induce farmers' field management changes. In 2012, it was suggested for the first time that farmers' motivation would be triggered if the demonstration was made that enhancing pollinator habitats, with a novel approach of farming with alternative pollinators, can increase yield and income. In 2013-2014, therefore, a 18month-pilot project was set on a participatory basis in Uzbekistan, to test this farming with alternative pollinators approach on field and orchard crops. The practicability and the potential of the approach were tested in collaboration with seven smallholders, two commercial farmers, and two schools. We analyzed the yield and insect diversity (pollinators, predators, and pests) of seven cucumber fields in the Parkent district and four orchards of sour cherry in the Boysun district in Uzbekistan. Here we show that the fields with enhanced habitats faced higher diversity of pollinators and predators, but less pests than control fields. Furthermore, the farming with alternative pollinators approach doubled the yield of sour cherry in 2014 and highly increased the income from cucumber in 2013. In 2014, however, a climatic disaster influenced the results on cucumber in Parkent district. Ultimately, 94% of the farmers were willing to enhance pollinator habitats after being informed of these higher-yield figures. If more projects confirm that farming with alternative pollinators creates an economically self-sustaining incentive for farmers to improve habitats, this approach could contribute considerably to global pollinator protection and food security.

Concern over exotic invasions is fueled in part by the observation that some exotic species appear to be more abundant and have stronger impacts on other species in their non-native ranges than in their native ranges. Past studies have addressed biogeographic differences in abundance, productivity, biomass, density and demography between plants in their native and non-native ranges, but despite widespread observations of biogeographic differences in impact these have been virtually untested. In a comparison of three sites in each range, we found that the abundance of Acroptilon repens in North America where it is invasive was almost twice that in Uzbekistan where it is native. However, this difference in abundance translated to far greater differences between regions in the apparent impacts of Acroptilon on native species. The biomass of native species in Acroptilon stands was 25-30 times lower in the non-native range than in the native range. Experimental addition of native species as seeds significantly increased the abundance of natives at one North American site, but the proportion of native biomass even with seed addition remained over an order of magnitude lower than that of native species in Acroptilon stands in Uzbekistan. Experimental disturbance had no longterm effect on Acroptilon abundance or impact in North America, but Acroptilon increased slightly in abundance after disturbance in Uzbekistan. In a longterm experiment in Uzbekistan, suppression of invertebrate herbivores and pathogens did not result in either consistent increases in Acroptilon biomass across years or declines in the biomass of other native species, as one might expect if the low impact of Acroptilon in the native range was due to its strong top-down regulation by natural enemies. Our local scale measurements do not represent all patterns of Acroptilon distribution and abundance that might exist at the scale of landscapes in either range, but they do suggest the possibility of fundamental biogeographic differences in the way a highly successful invader interacts with other species, differences that are not simply related to greater biomass or reduced topdown regulation of the invader in its non-native range.

An extended PCR method was established to rapidly identify and classify Bacillus thuringiensis strains containingcry (crystal protein) genes toxic to lepidopteran, coleopteran, and dipteran pests (Ben-Dov et al., Appl. Environ. Microbiol. 63:4883–4890, 1997). To optimize identification of all reported cry genes, this methodology needs a complete PCR set of primers. In the study reported here, a set of universal (Un9) and specific primers for multiplex rapid screening for all four known genes from the cry9 group was designed. PCR analyses were performed for cry9 genes on 16 standard strains and 215 field isolates of B. thuringiensis. Among the standard strains, only B. thuringiensis subsp. aizawaiHD-133, which harbors cry1 and cry2 genes, was positive with Un9 but negative to all four specific primers forcry9 genes. DNA of 22 field-collected isolates was also found to be positive with Un9. These isolates were classified into three cry9 profiles using specific primers; all of them harbor cry1 and cry2. This newly designed set of primers complements the existing PCR methodology for most currently known cry genes.

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