Chestnut blight fungus in Croatia: diversity of vegetative compatibility types,mating types and genetic variability of associated Cryphonectria hypovirus 1

In order to improve understanding of its diversity, 338 isolates of Cryphonectria parasitica, the causal agent of chestnut blight, were sampled from 10 chestnut populations throughout chestnut‐growing coastal and continental areas of Croatia. Eighteen vegetative compatibility (VC) types were identified. The VC type EU‐1 was the most widespread, comprising 42·9% of the isolates, followed by EU‐2 (21%) and EU‐12 (14·2%). In respect to the occurrence of the main VC types, the C. parasitica populations in Croatia combined features of both northwestern and southeastern European populations. Perithecia and mating‐type ratios of approximately 1 : 1 were found in all populations, suggesting that sexual reproduction of the fungus is common in Croatia. Natural hypovirulence was also evident in all populations, with incidence of hypovirus‐infected isolates ranging from 12·7% in Istria‐Buje to 66·6% in the continental part of the country. A total of 36 hypovirus‐infected isolates sampled throughout Croatia were analysed in ORF‐A and ORF‐B by RT‐PCR/RFLP analysis. All viral isolates belonged to the Italian subtype of Cryphonectria hypovirus 1 (CHV‐1) and were closely related to the isolates found in other European countries. The RFLP patterns found were also identical or similar to the patterns of three isolates collected in Croatia 22 years ago, suggesting a slow evolution of the hypovirus.     

Investigations on the role of the salmon louse,Lepeophtheirus salmonis (Caligidae), as a vector in the transmission of Aeromonas salmonicida subsp. salmonicida

A bacteria–parasite challenge model was used to study the role of sea lice, Lepeophtheirus salmonis (Copepoda), as a vector of Aeromonas salmonicida subsp. salmonicida. Three hypotheses were tested: (i) L. salmonis can acquire A. salmonicida subsp. salmonicida via water bath exposure; (ii) L. salmonis can acquire the bacteria via parasitizing infected Atlantic salmon, Salmo salar; and (iii) L. salmonis can transmit the bacteria to naïve Atlantic salmon via parasitism. Adult L. salmonis exposed to varying A. salmonicida subsp. salmonicida suspensions (10¹–10⁷ cells mL^?1) for 1.0, 3.0 or 6.0 h acquired the bacteria externally (12.5–100%) and internally (10.0–100%), with higher prevalences associated with the highest concentrations and exposures. After exposure to 10⁷ cells mL^?1, viable A. salmonicida subsp. salmonicida could be isolated from the external carapace of L. salmonis for 120 h. Lepeophtheirus salmonis also acquired the bacteria externally and internally from parasitizing infected fish. Bacterial transmission was observed only when L. salmonis had acquired the pathogen internally via feeding on ‘donor fish’ and then by parasitizing smaller (<50 g) ‘naive’ fish. Under specific experimental conditions, L. salmonis can transfer A. salmonicida subsp. salmonicida via parasitism; however, its role as a mechanical or biological vector was not defined.     

Effects of different patterns of feed restriction and insulin treatment during the luteal phase on reproductive, metabolic, and endocrine parameters in cyclic gilts

The objectives of the present study were 1) to study potential effects of previous nutritional treatment on developmental competence of early fertilized oocytes in vitro; 2) to study responses to insulin treatment during the period of feed restriction in the late luteal phase which has deleterious effects on subsequent fertility; and 3) to establish the metabolic and endocrine status of gilts during treatment and the subsequent periestrous period. Nineteen trios of littermate gilts were subjected to feed restriction during the first (RH) or second (HR) week of the estrous cycle. A second group of HR gilts received injections of long-acting insulin during their period of feed restriction (HR+I). Intensive sampling was performed in a subgroup of 23 animals on d 15 and 16 of the cycle for analyses of endocrine (gonadotropins and steroid hormones) and metabolic (insulin, IGF-I, leptin, total triiodothyronine [T3], and free T3) variables. Gilts were checked for estrus every 6 h, and time of ovulation was monitored by transcutaneous ultrasonography. Surgeries were performed 12 to 20 h after ovulation, and the early-fertilized oocytes recovered were cultured in vitro under standardized conditions. There was no treatment effect on the developmental competence of fertilized oocytes in vitro; however, ovulation rate was increased in HR+I gilts. No effect of treatment was observed on plasma leptin and IGF-I concentrations on d 15 and 16. However, HR+I gilts had higher (P < 0.05) postprandial insulin and lower (P < 0.05) postprandial total and free T3 on d 15. Plasma concentrations of LH, FSH, and progesterone on d 15 and 16 and plasma estradiol concentrations on d 16 were not affected by previous nutritional or insulin treatment. In the periestrous period, plasma concentrations of LH, FSH, and estradiol were higher (P < 0.05) in RH and HR+I, and the rise in plasma progesterone after the LH surge was lower (P < 0.05), than in HR gilts. No effect of treatment was observed on plasma concentrations of metabolic hormones, except on plasma leptin concentrations, which were higher (P < 0.05) at the time of the LH surge in RH gilts. These results suggest that feed restriction during the late luteal phase may have deleterious effects on ovarian function in the periestrous period, which may be counteracted by insulin.     

Low-density Ceratocystis polonica inoculation of Norway spruce (Picea abies) triggers accumulation of monoterpenes with antifungal properties

Among the most devastating pests of Norway spruce (Picea abies) are the European spruce bark beetle (Ips typographus) and the associated pathogenic blue-stain fungus Ceratocystis polonica. Following attack and colonization, the beetle and the fungus must cope with induced host chemical defenses, such as monoterpenes that are generally thought to be toxic to both symbionts. The goal of this study was to better understand the response of Norway spruce following C. polonica inoculation at low density that does not overwhelm the tree and to identify monoterpenes mobilized toward the fungus. We inoculated healthy mature trees and monitored monoterpene profiles 2, 3, and 5 months post-inoculation. We also exposed three different C. polonica strains to the most abundant or significantly up-regulated monoterpenes to determine differences in monoterpene toxicity and resistance among strains. Total monoterpene levels, including limonene, were increased at 2 and 3 months after inoculation and had dropped after 5 months. In in vitro assays, all monoterpenes were inhibitory to C. polonica. Limonene and β-pinene were the most potent inhibitors of fungal growth. The extent of inhibition varied between the three strains tested. These results showed a defense response of Norway spruce to C. polonica, in which limonene may play a critical role in inhibiting the spread of the fungus. We also showed that differences between strains of C. polonica must be taken into account when assessing the role of the fungus in this bark beetle–symbiont system.     

Separation, characterization, and quantitation of phenolic acids in a little-known blueberry (Vaccinium arctostaphylos L.) fruit by HPLC-MS

The aim of this study was the qualitative and quantitative determination of free, ester, glycoside, and ester-bound phenolic acids in the blueberry (Vaccinium arctostaphylos L.) fruit. A method for the determination of the profile of phenolic acids of four different phenolic fractions in the fruit was developed using high-performance liquid chromatography-mass spectrometry (HPLC-MS). Thirteen compounds (gallic, protocatechuic, p-hydroxybenzoic, m-hydroxybenzoic, gentisic, chlorogenic, p-coumaric, caffeic, ferulic, syringic, sinapic, salicylic, and trans-cinnamic acids) were identified and quantified in the berry. These experimental results showed that the predominant phenolic acid in the fruit of V. arctostaphylos is caffeic acid in free and insoluble ester-bound forms and p-coumaric acid in soluble ester and glycoside forms. Seven phenolic acids were identified as hydroxybenzoic acid derivatives (HBAs) and four as hydroxycinnamic acid derivatives (HCAs). Total content of HBAs and HCAs in the four phenolic fractions constituted 30.1 and 69.9% of the free, 27.9 and 72.1% of the ester, 24.7 and 75.3% of the glycoside, and 51.7 and 48.3% of the ester-bound forms, respectively. Total phenolics as the sum of individual phenolic acids identified is 698.5 ng/g of fresh weight (fw) for the free, 3399.2 ng/g of fw for the ester, 3522.1 ng/g of fw for the glycoside, and 3671.6 ng/g of fw for the ester-bound phenolic fractions. The present results were compared with reported levels of phenolic acids in the fruits of different Vaccinium species. These data suggest that the fruit can be considered as a potentially good dietary source of phenolic acids.     

Canopy reduction as a possible measure for adaptation of young Scots pine stand to insufficient precipitation in Central Europe

Climate change towards a warmer and dryer vegetation period may negatively impact growing conditions for Scots pine monocultures situated on dry, sandy soils in Central Europe. The purpose of the study was to evaluate the effect of thinning on precipitation throughfall in young Scots pine stands on typical pine sites. In 1992, observation of precipitation and throughfall started at the Tyniště research site (lowland of Eastern Bohemia) in a 7-year-old pine stand planted in rows at a stocking of ca 10,000 trees ha⁻¹. Throughfall was measured at weekly intervals during the growing season (April-September) by gauges randomly located in two treatments - variant 1C - Control without thinning and variant 2T - Thinned - and compared to precipitation at an open space outside of the canopy. The results demonstrated the positive effects of heavy low thinning (removal of 47% of the total number of trees and 31% of total pre-treatment basal area) on the water supply of young Scots pine stands. On the Thinned treatment, throughfall increased by 2-8% compared to Control plot. This positive effect persisted for six years after the first canopy reduction and the differences were significant for the first four years after thinning. After the second treatment (high thinning), throughfall on the Thinned treatment showed a nominal, but statistically insignificant increase. The likely reason for this result is that the application of a different type of thinning increased the variability of the canopy and, consequently, the effect of released crowns could not be detected.     

Food supplementation, nutritional intake of recipients and operational aspects: an integrated pilot nutrition initiative of BRAC

OBJECTIVES: To explore the nutritional quality of supplementary foods and additional energy consumption by the recipients in a pilot nutrition initiative of BRAC. DESIGN: In-depth interviews, observations during feeding at feeding centres, and laboratory analyses of supplementary foods for nutrient contents performed at the Institute of Nutrition and Food Science, University of Dhaka, Bangladesh. SETTING: Muktagacha thana (sub-district) in Mymensingh district, a rural area of Bangladesh. SUBJECTS: Pregnant and lactating mothers and children below 2 years of age. RESULTS: Analysis revealed that supplementary food, if taken completely, could provide daily energy equivalent to 752 kcal to a mother and 212 kcal to a child below 2 years of age. Mothers consumed about 75% of the food provided (approximately 564 kcal day(-1)). The food was shared mostly with young children and husbands. In-depth interview with mothers also suggested that they usually skipped breakfast if the food was given in the morning. The children liked the taste of food, and unless the child was sick or had some food before coming to the centre, she/he ate all the food provided. Although the main purpose of the project was to provide nutrition education, it was observed that activities at feeding centres were limited to food distribution with little time devoted to the communication of nutritional messages. CONCLUSIONS: Training should be given to service providers to communicate nutritional messages effectively as part of understanding the goal of the initiative. It is important to explore whether the regular diets of the recipients are replaced by the food supplementation or not.