Efficient elimination of sweetpotato little leaf phytoplasma from sweetpotato by cryotherapy of shoot tips

Shoot tips with 3–4 leaf primordia were excised from in vitro -grown sweetpotato plants ( Ipomoea batatas ) infected with little leaf phytoplasma ( Candidatus Phytoplasma aurantifolia) and subjected to cryotherapy. All plants regenerated from the cryo-treated shoot tips were free of phytoplasma, whereas shoot tip culture or dehydration of shoot tips without subsequent cryotherapy resulted in phytoplasma-free plants at a frequency of only 7–10%. Histological and ultrastructural studies with light and transmission electron microscopy, respectively, indicated that cryotherapy was lethal to all cells except those in the apical dome of the meristem and the two youngest leaf primordia. These surviving parts of the shoot tip contained vascular tissue and sieve elements, but electron microscopy showed no phytoplasma in them. In contrast, an abundance of phytoplasma was found in sieve elements located at the lower, non-surviving parts of the shoot tip 1·0 or 1·5 mm from the apical dome. In the greenhouse, the plants in which phytoplasmas were not detected were healthy-looking, grew vigorously and were readily distinguished from the infected plants that exhibited little leaf and chlorosis symptoms, proliferation of axillary shoots and roots, stunting, and heavily reduced number and size of storage roots. In this study efficient elimination of phytoplasma and production of pathogen-tested plant stocks were achieved with the novel cryotherapy-based approach. The proposed advantage of the technique is that it can be simultaneously used for long-term storage of plant germplasm and for production of pathogen-free plants.     

Estimating the germination dynamics of Plasmopara viticola oospores using hydro-thermal time

The effects of environmental conditions on the variability in germination dynamics of Plasmopara viticola oospores were studied from 1999 to 2003. The germination course was determined indirectly as the relative infection incidence (RII) occurring on grape leaf discs kept in contact with oospores sampled from a vineyard between March and July. The time elapsed between 1 January and the infection occurrence was expressed as physiological time, using four methods: (i) sums of daily temperatures > 8°C; (ii) hourly temperatures > 10°C; (iii) sums of hourly rates from a temperature-dependent function; or (iv) sums of these rates in hours with a rain or vapour pressure deficit ≤ 4·5 hPa (hydro-thermal time, HT). An equation of Gompertz in the form RII = exp[− a  · exp(− b  · HT)] produced an accurate fit for both separate years ( R ² = 0·97 to 0·99) and pooled data ( R ² = 0·89), as well as a good accuracy in cross-estimating new data ( r between observed and cross-estimated data were between 0·93 and 0·99, P  < 0·0001). It also accounted for a great part of the variability in oospore germination between years and both between and within sampling periods. Therefore, the equation of Gompertz (with a  = 15·9 ± 2·63 and b  = 0·653 ± 0·034) calculated over hydro-thermal time, a physiological time accounting for the effects of both temperature and moisture, produced a consistent modelling of the general relationships between the germination dynamics of a population of P. viticola oospores and weather conditions. It represents the relative density of the seasonal oospores that should have produced sporangia when they have experienced favourable conditions for germination.     

Genetic diversity of sulfonylurea-resistant and -susceptible Monochoria vaginalis populations in Japan

Monochoria vaginalis is one of the most serious weeds of rice fields in Asia. The species is predominantly selfing. To reveal the potential for multiple mutational events, outcrossing and gene flow in the sulfonylurea‐resistant (SU‐R) M. vaginalis populations, we investigated (i) if each SU‐R population was a single SU‐R biotype or a mixture of several SU‐R biotypes using restriction analysis or direct sequencing of acetolacatate synthase (ALS) genes and (ii) genetic diversity of SU‐R and ‐susceptible (S) populations using amplified fragment length polymorphism (AFLP) analysis. Nineteen or 20 individuals were sampled from four SU‐R and five SU‐S populations respectively. Amino acid substitutions conferring resistance in the SU‐R populations were Pro₁₉₇Ser in the ALS1 or ALS3, or Asp₃₇₆Glu in the ALS1 and each SU‐R population was composed of a single SU‐R biotype. In cluster analysis each SU‐R individual formed a cluster, whereas the individuals from a SU‐S population belonged to different clusters. Some SU‐R populations showed polymorphic AFLP loci. The results indicated that these SU‐R biotypes emerged from a single mutational event and any gene flow of SU‐R genes from adjacent populations did not occur. A low level of outcrossing and recombinations of SU‐R genes occurred within some SU‐R populations of M. vaginalis.     

Significance of weed hosts for Potato virus Y protection in Syria

A survey of Potato virus Y (PVY) was carried out on weeds growing in and around potato fields in Syria during the autumn growing seasons of 2002, 2004 and 2006. A total of 59 samples of eight weed species and three tobacco ( Nicotiana tabacum ) samples were tested by ELISA using PVY antisera. Among them 15 samples belonged to Solanum nigrum (7 samples), Physalis sp. (6 samples) and tobacco (2 samples) were PVY infected. This suggests that weed hosts and neighbouring tobacco fields are natural reservoirs of PVY in Syria. According to their biological, serological and molecular characteristics, the majority of weed PVY isolates belonged to the PVY^SYR variant indicating a possible correlation between the high incidence of PVY^SYR in potato and weed hosts. This is the first report on the occurrence and characterization of weed PVY isolates from Syria.     

A yellows disease system with differing principal host plants for the obligatory pathogen and its vector

A stolbur‐type phytoplasma is the putative pathogen of grapevine yellows disease that causes economic damage to vineyards in most growing areas around the world. The pathogen is known to be transmitted to vines by two planthoppers, Hyalesthes obsoletus and Reptalus panzer; the latter is found in Europe but has not yet been observed in Israel. The establishment of a vector–pathogen–plant relationship requires that the pathogen and the vector meet on a shared host plant. This does not happen in the ecosystem examined here, where two different principal host plants for the obligate pathogen and its vector exist: the pathogen is established on vines, while its vector, H. obsoletus, develops on Vitex agnus‐castus. The present study verified that: (i) the vector cannot complete its life cycle on vines; (ii) V. agnus‐castus does not grow in the immediate vicinity of vines, and does not harbour the pathogen; and (iii) the pathogen is not vertically transmitted from mother to offspring. Moreover, in a thorough search of plants in vine growing areas, no other plants were found that host both the vector and the pathogen. However, it was found that the planthopper can acquire the phytoplasma from infected vines. Nonetheless, this does not prove the ability of the planthopper to further transmit the pathogen to vines and does not explain the presence of the vector on the non‐preferred vines. Thus, the enigma of the pathogen–vector–host triangle in this system remains unresolved.     

Natural tolerance of Cuscuta campestris to herbicides inhibiting amino acid biosynthesis

The response of Cuscuta campestris Yuncker, a non‐specific above‐ground holoparasite, to amino acid biosynthesis inhibitor (AABI) herbicides, was compared with other resistant and sensitive plants in dose–response assays carried out in Petri dishes. Cuscuta campestris was found to be much more resistant to all AABI herbicides tested. The I₅₀ value of C. campestris growth inhibition by glyphosate was eightfold higher than that of transgenic, glyphosate‐resistant cotton (RR‐cotton). The I₅₀ value for C. campestris shoot growth inhibition by sulfometuron was above 500 μM, whereas that of sorghum roots was only 0.004 μM. Cuscuta campestris exposed to glyphosate gradually accumulated shikimate, confirming herbicide penetration into the parasite and interaction with an active form of the target enzyme of the herbicide, 5‐enolpyruvylshikimate‐3‐phosphate synthase. More than half of the C. campestris plants associated with transgenic, glyphosate‐resistant sugarbeet (RR‐sugarbeet) treated with glyphosate or with transgenic, sulfometuron‐resistant tomato (SuR‐tomato) treated with sulfometuron recovered and resumed regular growth 20–30 days after treatment. New healthy stems developed, followed by normal flowering and seed setting. The results of the current study demonstrate the unique capacity of C. campestris to tolerate high rates of AABI. The mechanism of this phenomenon is yet to be elucidated.     

Detection, identification and significance of phytoplasmas in grasses in northern Australia

Sugarcane yields have been severely reduced by white leaf and grassy shoot phytoplasma diseases in many parts of Asia. Australian sugarcane crops are not known to be affected by these diseases, but plant pathogenic phytoplasmas found in other introduced and native grasses in northern Australia could pose a serious threat to the Australian sugarcane industry. To further evaluate this threat, leaves from plants of 20 grass species, with and without symptoms, were collected during field surveys in northern Australia and tested to determine whether phytoplasmas were present and whether symptoms were reliable indicators of phytoplasma presence. Molecular tools were used to detect and characterize phytoplasmas. Four different phytoplasmas were found in seven grass species known to grow near healthy sugarcane crops. All the phytoplasmas were closely related to sugarcane white leaf phytoplasma (SCWL), one of the phytoplasmas that causes disease in sugarcane in Asia. Four of the host plant species and two of the phytoplasmas were new records. The relationship between symptoms and phytoplasma presence was poor. Because some plants with symptoms tested negative for phytoplasmas, a series of surveys was carried out in which flowers, leaves, roots and stems of two known host plant species, Whiteochloa cymbiformis and Sorghum stipoideum, were tested separately on nine occasions during two wet seasons. This was done to investigate the distribution of phytoplasmas within plants over time. Results showed that spatial and temporal variation of phytoplasmas occurred in these two host plant species. Hence, evaluation of disease distribution within a region requires repeated testing of all plant parts from plants without symptoms, as well as those with symptoms. To date, there is no report of a vector capable of transmitting to Australian sugarcane the phytoplasmas found in grasses in this study. If one is present, or occurs in the future, then native and introduced grasses could constitute a large reservoir of phytoplasma for vectors to draw on. This work provides an early warning for the sugarcane industry that the potential for infection exists.     

Predicting effective doses for the joint action of two fungicide applications

A function was derived to predict fungicide efficacy when more than one application of a single active ingredient is made to a crop, given parameters describing the dose–response curves of the component single-spray applications. In the function, a second application is considered to act on that proportion of the total pathogen population which was uncontrollable at the time of the first application (represented by the lower asymptote of the dose–response curve for the first treatment), plus any additional part of the population which survived the first application as a result of a finite dose being applied. Data to estimate the single-spray dose–response curve parameters and validate predictions of two-spray programme efficacy were obtained from separate subsets of treatments in four field experiments. A systemic fungicide spray was applied to wheat at a range of doses, at one or both of two times (t₁ and t₂), in all dose combinations. Observed values of the area under the disease progress curve (AUDPC) for septoria leaf blotch (Mycosphaerella graminicola) were used to construct response surfaces of dose at t₁ by dose at t₂ for each culm leaf layer. Parameters were estimated from single-spray and zero-dose treatment data only. The model predicted a high proportion (R² = 71–95%) of the variation in efficacy of the two-spray programmes. AUDPC isobols showed that the dose required at t₂ was inversely related to the dose at t₁, but the slope of the relationship varied with the relative timings of t₁ and t₂ in relation to culm leaf emergence. Isobols were curved, so the effective dose – the total dose required to achieve a given level of disease suppression – was lower when administered as two applications.