Ken J. Devine

The influence of in-egg mortality and spontaneous hatching on the decline of Globodera rostochiensis during crop rotation in the absence of the host potato crop in the field

The decline of Globodera rostochiensis (Woll.) populations in two fields was 57% during the first and 40.3% during the second year of crop rotation in the absence of a potato crop in Co. Cork, Ireland. The decline was found to be due to both spontaneous hatch and in-egg mortality. Spontaneous hatch accounted for between 75.8 and 80.2% of the decline experienced during rotation. In-egg mortality was found to be responsible for a reduction of approximately 10% in the total number of viable eggs in both fields, and was positively correlated with soil temperature in the field. The number of culturable bacteria and the composition of the bacterial microflora in the cysts was found to change with increasing soil temperatures. The increase in cellulase-positive isolates reflected the effect of temperature on egg degradation. Der Einfluss von Absterben im Ei und spontanem Schlupfen auf die Abnahme von Globodera rostochiensis im Feld im Laufe des Fruchtwechsels bei Abwesenheit des Wirtes Kartoffel - In Co. Cork, Irland, betrug die Abnahme der Populationen von Globodera rostochiensis (Woll.) in zwei Feldern in einer Fruchtfolge ohne Kartoffeln im ersten Jahr 57% und im zweiten Jahr 40,3%. Es wurde festgestellt, dass die Abnahme durch spontanes Schlupfen und durch Absterben im Ei verursacht wurde. Spontanes Schlupfen war fur 75,8 und 80,2% des wahrend des Fruchtwechsels beobachteten Ruckganges verantwortlich. Etwa 10% der Gesamtzahl infektionsfahiger Eier in den beiden Feldern gingen durch Absterben im Ei zugrunde. Dieser Ruckgang war positiv korreliert mit der Bodentemperatur im Feld. Der Anteil der kulturfahigen Bakterien und die Zusammensetzung der Bakterienflora in den Zysten nahmen mit steigenden Bodentemperaturen zu. Der Anstieg der zellulase-positiven Isolate spiegelte die Wirkung der Temperatur auf die Zersetzung der Eier wider.

Lipidic dispersion to reduce food dependent oral bioavailability of fenofibrate: In vitro, in vivo and in silico assessments.

Novel formulations that overcome the solubility limitations of poorly water soluble drugs (PWSD) are becoming ever more critical to a drug development process inundated with these compounds. There is a clear need for developing bio-enabling formulation approaches to improve oral bioavailability for PWSD, but also to establish a range of predictive in vitro and in silico biopharmaceutics based tools for guiding formulation design and forecasting in vivo effects. The dual aim of this study was to examine the potential for a novel lipid based formulation, termed a lipidic dispersion, to enhance fasted state oral bioavailability of fenofibrate, while also assessing the predictive ability of biorelevant in vitro and in silico testing. Formulation as a lipidic dispersion improved both dissolution and solubilisation of fenofibrate through a combination of altered solid state characteristics and incorporation of solubilising lipidic excipients. These changes resulted in an increased rate of absorption and increased maximal plasma concentrations compared to a commercial, micronised product (Lipantil® Micro) in a pig model. Combination of biorelevant in vitro measurements with in silico physiologically based pharmacokinetic (PBPK) modelling resulted in an accurate prediction of formulation performance and forecasts a reduction in food effects on fenofibrate bioavailability through maximising its fasted state dissolution.

Investigations into the chemoattraction of the potato cyst nematodes Globodera rostochiensis and G. pallida towards fractionated potato root leachate

The behaviour of stimulated second stage juveniles (J2) (i.e., hatched in root leachate from potato cv. Cara) and unstimulated J2 (spontaneously hatched in water) of Globodera rostochiensis and G. pallida in response to fractionated and unfractionated potato root leachate (PRL) was investigated in attraction assays. In PRL, fractionated by combined ion-exchange-gel permeation chromatography on Sephadex G-10, three classes of semiochemicals with activity towards J2 were distinguished: i) chemoattractants; ii) chemostats, and iii) chemorepellents. The motility of PRL-hatched G. rostochiensis J2 in one fraction (12) at 10 days after their removal from the root leachate was significantly greater than that of water-hatched J2 apparently due to sensitisation of PRL-hatched J2. PRL-hatched J2 of G. pallida were attracted to different fractions than those of G. rostochiensis, whereas the water-hatched J2 from the two species were attracted to common fractions, indicating that sensitisation by exposure to PRL was species selective. The attraction of PRL-hatched PCN J2 to unfractionated PRL appeared to be dependent on the ratio of chemoattractant to chemorepellent semiochemicals in the leachate. For both species there was no detectable correlation between hatching activity and either attractiveness of root leachates from 12 potato genotypes or chemoattraction in PRL fractions, indicating that hatching factors were not active chemoattractants.

Effects of hatching factors on potato cyst nematode hatch and in-egg mortality in soil and in vitro

The exogenous application to three Globodera pallida-infested fields of tomato root leachate (TRL) containing hatching factors increased nematode hatch and in-egg mortality, particularly in a highly organic soil, and in a sandy but not in a clay soil. The most active concentrations of TRL (7.5-12.5 mg m-2) resulted in a reduction of between 69 and 79% in the number of viable eggs per cyst recovered 12 weeks after TRL application. At high hatching factor concentrations, supra-optimal inhibition of both hatch and in-egg mortality was observed; generally, hatch and in-egg mortality exhibited similar dosage-responses to TRL. A significant TRL dosage-hatch response was observed at 4 weeks after TRL application only in the sandy soil. In vitro, a G. pallida population exhibiting moderate hatch gave similar hatching and in-egg mortality responses as in the field experiment. The response of the PCN populations in vitro was found to be dependent on the physiological state of the egg/juvenile complex, with egg populations in diapause responding to the presence of natural and artificial hatching factors by exhibiting increased in-egg mortality but not increased hatch. The results are discussed in relation to novel G. pallida control measures.

Studies on the effect of mycorrhization of potato roots on the hatching activity of potato root leachate towards the potato cyst nematodes, Globodera pallida and G. rostochiensis

Successful mycorrhization of potato plants cv. Golden Wonder was achieved with three commercial preparations of arbuscular mycorrhizal fungi (AMF): Vaminoc (mixed-isolate inoculum) and two of its components, Glomus intraradices and Glomus mosseae. Potato cyst nematode hatching assays were conducted on the potato root leachate (PRL) produced by inoculated and non-inoculated potato plants to examine the effect of AMF inoculation on the hatching response of the two PCN species, Globodera rostochiensis and G. pallida. The overall hatch response of G. rostochiensis to the potato root leachate was greater than G. pallida. Root leachates from Vaminoc- and G. mosseae-inoculated plants were found to stimulate the hatch of G. pallida in the first 3 weeks after shoot emergence. Fractionation of root leachates with standardised carbon content by Sephadex G-10 chromatography revealed multiple AMF effects on hatching factor (HF) production. Root leachates from Vaminoc-inoculated plants contained markedly more G. pallida-active HF than all other treatments; by contrast, PRL from the three AMF treatments exhibited little variation in the quantity of G. rostochiensis-active HF produced. Several HF were PCN species-specific or species-selective, with those resolved from the G. intraradices and G. mosseae PRL profiles exhibiting an apparent preference for G. rostochiensis rather than G. pallida. Mycorrhization also significantly increased the root dry weight of plants.

In vitro studies on the relative availability and mobility in soil of natural hatching factors for the potato cyst nematodes, Globodera rostochiensis and G. pallida

The fractionation of root leachates from tomato and potato on a Sephadex G-10 column revealed many similarities between the elution profiles of hatching activities towards potato cyst nematodes (PCN). When aliquots of either tomato or potato root leachate were mixed with different soil samples, hatching factors exhibiting G. pallida-selective hatching activity showed less affinity for the soil matrix than did those selective for G. rostochiensis . This was confirmed by the earlier elution of G. pallida-selective hatching factors from columns in which soil was used as the solid phase in low pressure liquid chromatography. The selectivity of the earliest-eluting hatching factors towards G. pallida, relative to G. rostochiensis, was found to increase significantly as the percentage soil organic matter content increased.

Comparison of the production and mobility of hatching activity towards the potato cyst nematodes, Globodera rostochiensis and G. pallida within soil planted with a host potato crop

Soil core samples were taken from a commercial potato field before emergence of the potato crop, during crop development and after harvesting. Leachates from the cores were analysed in an in vitro hatching assay in the laboratory for activity towards the potato cyst nematodes (PCN), Globodera rostochiensis and G. pallida. Hatching activity in the soil increased rapidly after plant emergence and peaked between 2 and 5 weeks after emergence; thereafter, at about the onset of flowering, hatching factor activity decreased markedly. The soil cores collected in the first 2 weeks after plant emergence contained significantly greater hatching activity towards G. pallida than G. rostochiensis, indicating that G. pallida-selective hatching factors were produced by the host crop earlier than G. rostochiensis-selective hatching factors. Soil cores collected from the potato ridge and the furrows at various depths and distances from the host plants differed in the distribution of G. pallida-selective and G. rostochiensis-selective hatching activity within the soil profile. Globodera pallida-selective hatching activity had greater vertical and horizontal mobility in the soil profile and elicited significantly greater hatching responses than G. rostochiensis beyond the region of the rhizosphere. Significant levels of hatching factor activity could be detected in the field 90 days after harvesting of the potato crop.

Potato cyst nematode hatching activity and hatching factors in inter-specific Solanum hybrids

In studies using three sibling F1 clones from each of five crosses between Solanum tuberosum and different wild tuberbearing Solanum species, root leachate hatching activity towards each of the two potato cyst nematode (PCN) species, Globodera rostochiensis and G. pallida , segregated independently. Sephadex G-10 fractionation of the leachates of S. sucrense -hybrid clones revealed differences between clones in the hatching factor elution profiles for the two PCN species. Analysis of individual hatching factors indicated examples both of selectivity (both PCN species hatched in response to a particular hatching factor, but at different levels of hatching response) and of specificity (only one PCN species hatched in response to a particular hatching factor) of hatching factors. The hatching factor profiles of the S. sucrense -hybrids indicated segregation of hatching factors from the parents, but also detected novel factors unique to specific hybrid clones. Total hatching activity of the root leachate of the S. sucrense clones was positively correlated with the proportion of S. tuberosum -derived hatching factors.

Broadband Acoustic Resonance Dissolution Spectroscopy (BARDS): A Rapid Test for Enteric Coating Thickness and Integrity of Controlled Release Pellet Formulations.

Graphical abstract Figure. No Caption available. Abstract There are no rapid dissolution based tests for determining coating thickness, integrity and drug concentration in controlled release pellets either during production or post‐production. The manufacture of pellets requires several coating steps depending on the formulation. The sub‐coating and enteric coating steps typically take up to six hours each followed by additional drying steps. Post production regulatory dissolution testing also takes up to six hours to determine if the batch can be released for commercial sale. The thickness of the enteric coating is a key factor that determines the release rate of the drug in the gastro‐intestinal tract. Also, the amount of drug per unit mass decreases with increasing thickness of the enteric coating. In this study, the coating process is tracked from start to finish on an hourly basis by taking samples of pellets during production and testing those using BARDS (Broadband Acoustic Resonance Dissolution Spectroscopy). BARDS offers a rapid approach to characterising enteric coatings with measurements based on reproducible changes in the compressibility of a solvent due to the evolution of air during dissolution. This is monitored acoustically via associated changes in the frequency of induced acoustic resonances. A steady state acoustic lag time is associated with the disintegration of the enteric coatings in basic solution. This lag time is pH dependent and is indicative of the rate at which the coating layer dissolves. BARDS represents a possible future surrogate test for conventional USP dissolution testing as its data correlates directly with the thickness of the enteric coating, its integrity and also with the drug loading as validated by HPLC.