Identification of ALS inhibitor‐resistant Amaranthus biotypes using polymerase chain reaction amplification of specific alleles

Summary Polymerase chain reaction (PCR) amplification of specific alleles (PASA) was adapted as a molecular marker‐based method for the rapid detection of point mutations in Amaranthus retroflexus and Amaranthus rudis leading to ALS inhibitor resistance. Two pairs of primers were designed for the specific amplification of alleles of the ALS gene of susceptible and resistant biotypes. The allele‐specific primer matched the desired allele, but mismatched the different allele at its 3′ end. Differentiation was carried out by comparison of the amplified DNA fragments in gel electrophoresis after PASA‐PCR. In A. rudis, differentiation was possible with one PCR and genomic DNA as probe. A ‘nested’ PCR was necessary for the differentiation of sensitive and resistant A. retroflexus. PASA is useful for the identification of resistant weed biotypes and also as a monitoring tool to map resistance occurrence and distribution. Advantages include the fast and clear separation of those plants with and without mutations at an early stage of development, its easy and consistent performance and quick results compared with existing resistance detection tests. These advantages, when combined with management strategies, enable further activities to reduce herbicide resistance.     

Orobanche species and population discrimination using intersimple sequence repeat (ISSR)

Summary Orobanche species are commonly identified using morphological characteristics. In many cases, the distinction of closely related species is difficult, and a molecular tool is more suitable to differentiate them. In this study, genomic polymorphism between morphologically distinct species was investigated through amplification by polymerase chain reaction (PCR) of intersimple sequence repeat (ISSR) regions. Five primers were used to study genetic variation in the morphologically distinct species O. hederae and O. amethystea, as well as the closely related species O. cernua and O. cumana. For the first two species, all the primers detected genetic polymorphism. Anchored primers allowed the identification of more specific molecular markers than non‐anchored tri‐ and tetranucleotide primers. Genetic polymorphism was investigated among three O. hederae populations using the two types of primer. One non‐anchored and two anchored primers detected intraspecific variation, which was not correlated with the geographical location of those populations. The primer (GATA)₄ detected polymorphism between five specimens each of O. cernua and O. cumana species collected from different countries, permitting these two closely related species to be clearly differentiated. This study demonstrated that ISSR markers can be highly reliable for precise identification of Orobanche species.     

A leakiness index for assessing landscape function using remote sensing

The cover, number, size, shape, spatial arrangement and orientation of vegetation patches are attributes that have been used to indicate how well landscapes function to retain, not ‘leak’, vital system resources such as rainwater and soil. We derived and tested a directional leakiness index (DLI) for this resource retention function. We used simulated landscape maps where resource flows over map surfaces were directional and where landscape patch attributes were known. Although DLI was most strongly related to patch cover, it also logically related to patch number, size, shape, arrangement and orientation. If the direction of resource flow is multi-directional, a variant of DLI, the multi-directional leakiness index (MDLI) can be used. The utility of DLI and MDLI was demonstrated by applying these indices to three Australian savanna landscapes differing in their remotely sensed vegetation patch attributes. These leakiness indices clearly positioned these three landscapes along a function-dysfunction continuum, where dysfunctional landscapes are leaky (poorly retain resources). This revised version was published online in July 2006 with corrections to the Cover Date.     

Spatial dependence and the relationship of soil organic carbon and soil moisture in the Luquillo Experimental Forest,Puerto Rico

We used geo-spatial statistical techniques to examine the spatial variation and relationship of soil organic carbon (SOC) and soil moisture (SM) in the Luquillo Experimental Forest (LEF), Puerto Rico, in order to test the hypothesis that mountainous terrain introduces spatial autocorrelation and crosscorrelation in ecosystem and soil properties. Soil samples (n = 100) were collected from the LEF in the summer of 1998 and analyzed for SOC, SM, and bulk density (BD). A global positioning system was used to georeference the location of each sampling site. At each site, elevation, slope and aspect were recorded. We calculated the isotropic and anisotropic semivariograms of soil and topographic properties, as well as the cross-variograms between SOC and SM, and between SOC and elevation. Then we used four models (random, linear, spherical and wave/hole) to test the semi-variances of SOC, SM, BD, elevation, slope and aspect for spatial dependence. Our results indicate that all the studied properties except slope angle exhibit spatial dependence within the scale of sampling (200 – 1000 m sampling interval). The spatially structured variance (the variance due to the location of sampling sites) accounted for a large proportion of the sample variance for elevation (99%), BD (90%), SOC (68%), aspect (56%) and SM (44%). The ranges of spatial dependence (the distances within which parameters are spatially dependent) for aspect, SOC, elevation, SM, and BD were 9810 m, 3070 m, 1120 m, 930 m and 430 m, respectively. Cross correlograms indicate that SOC varies closely with elevation and SM depending on the distances between samples. The correlation can shift from positive to negative as the separation distance increases. Larger ranges of spatial dependence of SOC, aspect and elevation indicate that the distribution of SOC in the LEF is determined by a combination of biotic (e.g., litterfall) and abiotic factors (e.g., microclimate and topographic features) related to elevation and aspect. This demonstrates the importance of both elevation and topographic gradients in controlling climate, vegetation distribution and soil properties as well as the associated biogeochemical processes in the LEF.This revised version was published online in May 2005 with corrections to the Cover Date.     

Simulating evolution of glyphosate resistance in Lolium rigidum II: past, present and future glyphosate use in Australian cropping

Glyphosate is a key component of weed control strategies in Australia and worldwide. Despite widespread and frequent use, evolved resistance to glyphosate is rare. A herbicide resistance model, parameterized for Lolium rigidum has been used to perform a number of simulations to compare predicted rates of evolution of glyphosate resistance under past, present and projected future use strategies. In a 30‐year wheat, lupin, wheat, oilseed rape crop rotation with minimum tillage (100% shallow depth soil disturbance at sowing) and annual use of glyphosate pre‐sowing, L. rigidum control was sustainable with no predicted glyphosate resistance. When the crop establishment system was changed to annual no‐tillage (15% soil disturbance at sowing), glyphosate resistance was predicted in 90% of populations, with resistance becoming apparent after between 10 and 18 years when sowing was delayed. Resistance was predicted in 20% of populations after 25–30 years with early sowing. Risks of glyphosate resistance could be reduced by rotating between no‐tillage and minimum‐tillage establishment systems, or by rotating between glyphosate and paraquat for pre‐sowing weed control. The double knockdown strategy (sequential full rate applications of glyphosate and paraquat) reduced risks of glyphosate and paraquat resistance to <2%. Introduction of glyphosate‐resistant oilseed rape significantly increased predicted risks of glyphosate resistance in no‐tillage systems even when the double knockdown was practised. These increased risks could be offset by high crop sowing rates and weed seed collection at harvest. When no selective herbicides were available in wheat crops, the introduction of glyphosate‐resistant oilseed rape necessitated a return to a minimum‐tillage crop establishment system.     

Identification of a major gene (Mex-1) from Coffea canephora conferring resistance to Meloidogyne exigua in Coffea arabica

Among the most damaging root-knot nematode species, Meloidogyne exigua is especially common in Latin America and constitutes a major agronomic constraint in all major coffee-growing ( Coffea arabica ) areas. Growing nematode-resistant coffee represents the most promising option for control of the pest. The present study aimed to determine the mode of inheritance of the M. exigua resistance transferred into C. arabica from a related species, Coffea canephora , and to identify associated molecular markers. Segregation data analysis of F ₂ progeny derived from a cross between the resistant introgression line T5296 and the susceptible accession Et6 showed that the resistance to M. exigua is controlled by a simply inherited major gene (designated the Mex -1 locus). The gall index distribution exhibited by the F ₂ individuals suggested incomplete dominant expression. Fourteen AFLP markers were found associated with the resistance to M. exigua and a localized genetic map of the chromosome segment carrying Mex -1 was constructed. Furthermore, the association of the identified AFLP markers with Mex -1 was confirmed by analysis of a set of genotypes involving 28 introgression Arabica lines either resistant or susceptible to M. exigua in field conditions. These results represent an important starting point to enhance backcross breeding programmes and to perform an early selection of resistant seedlings.