Epigallocatechin‐3‐gallate (EGCG) and green tea polyphenols do not improve stallion semen parameters during cooling at 4°C

Stallion semen storage for artificial insemination is mainly based on liquid cooled storage. In many stallions this technique maintains sperm quality for an extended period of time (24–72 hr) at 7°C. While this technique is commonly used in the horse industry, there can be a decline in fertility in some stallions, due to an inability of their sperm to tolerate the cool storage process. The aim of the present work was to evaluate the effect of two natural antioxidants (epigallocatechin‐3‐gallate (EGCG) at 20, 60 and 120 μm and green tea polyphenols, and p at .001, .01 and .1 mg/ml) on some sperm parameters (sperm motility, viability/acrosome integrity and DNA quality) in extended semen immediately after its collection (T0) and after 2, 6, 24 and 48 hr of cool storage. Two ejaculates from three trotter stallions were analysed after 48 hr of storage at 4°C. No beneficial effect on the analysed parameters was observed: the two antioxidants were not able to improve sperm quality after 48 hr of storage. These results are in agreement with previous findings on the effect of different antioxidants reported by other researches, who have demonstrated that stallion semen keeps good antioxidant capacity after dilution for 24 hr. In conclusion, the positive effect exerted by antioxidant molecules in other species is not confirmed in the equine one.     

Characterization of Neofabraea vagabunda isolates causing apple bull's eye rot in Italy (Emilia‐Romagna region)

Neofabraea vagabunda is the prevalent cause of bull's eye rot, one of the main postharvest diseases of apple, in many producing areas, but its biology has not been studied in detail. The molecular identification, by DNA sequencing of the β‐tubulin region, of 41 isolates collected from apples showing bull's eye rot in the Emilia‐Romagna region confirmed N. vagabunda as the main species in Italy. A biological and morphological characterization of N. vagabunda isolates was performed in vitro. Assays at temperatures ranging from 0 to 30 °C carried out on 10 isolates demonstrated: (i) a marked influence of temperature on colony morphology, conidial production, conidial size and mycelial growth, showing the cold‐tolerant character of N. vagabunda; and (ii) that culture at 15 °C on tomato agar (TA) for 14 days is a rapid and reliable method to favour pathogen conidial production. Trials performed on 38 isolates using these incubation conditions recorded the presence of two N. vagabunda morphotypes, differing for colony morphology, conidial size, conidiomata formation and temperature requirement. The alkalizing ability of the pathogen during growth on TA was also demonstrated for the first time. The pathogenicity of 25 N. vagabunda isolates was proved in vivo on artificially infected Cripps Pink apples. A pH increase was also recorded in apple tissue infected by N. vagabunda isolates (on average 0.2 and 0.3 units of pH after 60 and 120 days of incubation, respectively), suggesting that the N. vagabunda transition from quiescence to necrotrophic colonization in apples could involve the secretion of alkalizing compounds.     

Evaluation of the mineralizable organic nitrogen in soil using an immunological technique

Phosphate buffer-extractable organic nitrogen (PEON), which can be extracted from soil with a 67-mM neutral phosphate buffer solution, is considered to be a major source of mineralizable nitrogen (N) that is ubiquitous in soil. We have developed a Western blot analysis and enzyme-linked immune sorbent assay (ELISA) method for determining PEON. First, anti-PEON antibody (IgG) from a rabbit was produced using purified PEON from a volcanic ash soil. PEON was detected as a ladder-like staining pattern in the high-molecular-weight region and as a single band under 14 kDa in Western blot analysis using the anti-PEON IgG. In the ELISA assay, the anti-PEON IgG could specifically react with PEON irrespective of soil type and management. Furthermore, the absorbance at 450 nm in ELISA was highly correlated to the concentration of organic N (N_org) in phosphate extracts of different soils. We propose the immune assay using the anti-PEON IgG as a possible novel method for evaluating available N in soil.     

Invertebrate communities (Annelida and epigeic fauna) in three types of Estonian cultivated soils

The abundance and diversity of invertebrate communities (annelids and epigeic fauna) in three types of cultivated soils were studied. Soil biota communities in the three most widespread soil types in Estonia (Calcaric Regosols, Calcaric Cambisols and Stagnic Luvisols) are influenced by environmental conditions, the factors connected to soil texture including moisture, organic matter content and pH being the most essential, and by the intensity of agricultural practice. Potentially high biological activity and low intensity of agricultural human activity of Calcaric Regosols occurs in parameters of communities of organisms not sensitive to soil which dries off, i.e. epigeic fauna living on the soil surface and preferring dry and warm habitat; temporarily dried off soil is not a suitable habitat for Oligochaeta. Both groups of Oligochaeta (earthworms, enchytraeids) appear to prefer Calcaric Cambisols where soil moisture conditions are more stable. The abundance of invertebrate communities is the highest and the diversity is the lowest in Stagnic Luvisols. Some trends occurred in community characteristics along the soil surface following a hypothetical gradient; the number of carabids per trap and diversity of spiders decreased from the edge to the centre of the field. The results presented here on spatial variability in distribution of soil organisms are preliminary.     

Chilling-induced oxidative stress and antioxidant responses in mume (Prunus mume) fruit during low temperature storage

Mume (Prunus mume Sieb. et Zucc.) fruit are harvested and consumed at the mature green stage and have a short storage life at ambient temperature. While cold temperature extends their storage life, improper refrigeration causes severe chilling injury (CI), with fruit suffering more severe CI at of 5–6 °C than at 1 °C. The objective of this research was to determine the involvement of reactive oxygen species (ROS) and antioxidant systems in fruit under chilling stress. ‘Nankou’ fruit were stored at 1 °C or 6 °C for 15 days. Hydrogen peroxide, a preventive ROS, decreased at a slower rate at 6 °C than 1 °C during storage. Malondialdehyde (MDA), an indicator of lipid peroxidation caused by ROS, increased during storage and the contents were higher in fruit stored at 6 °C than at 1 °C. On the other hand, fruit stored at 6 °C had a lower total antioxidant capacity (TAC) and lower activities of antioxidant-related enzymes including superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) than at 1 °C. These results indicate that the fruit at 6 °C had more oxidative stress; thus the fruit had more severe CI symptoms than at 1 °C.     

Anti-Brucella antibodies in seals at coastal locations of Hokkaido,Japan, with focus on life stages

The enzyme-linked immunosorbent assay (ELISA) was applied to detect antibodies against Brucella abortus in serum samples from four seal species at nine coastal locations of Hokkaido, Japan. These antibodies were detected in 27% (32/118) of Western Pacific harbor seals (Phoca vitulina stejnegeri) at Cape Erimo. The antibodies were observed in spotted seals (P. largha) in one out of six at Nemuro, in two out of three at Rebun Island, in one out of two at Bakkai, and in examined one at Soya. They were also found in respective examined one ribbon seal (Histriophoca fasciata) and one ringed seal (Pusa hispida) at Akkeshi. Harbor seals that tested positive were mostly yearlings (35%, 20/57) and juveniles (45%, 10/22), while only one pup (1/13) and one subadult (1/5) tested positive with low titers of the antibody; no antibodies were observed in adults (n=21). These results suggest that Brucella mainly infected harbor seals from the environment while weaning, and the bacteria were cleared during the early life stage of the seals. In spotted seals, however, antibodies were also detected in adults, suggesting that spotted seals could become infected with Brucella even as adults. It is also possible that a different, more persistent strain of Brucella may have infected the spotted seals.     

Genital mycobacteriosis caused by Mycobacterium marinum detected in two captive sharks by peptide nucleic acid–fluorescence in situ hybridization

Mycobacterium marinum is a prevalent nontuberculous mycobacterium (NTM)-infecting teleosts. Conversely, little is known about mycobacteriosis in elasmobranchs, and M. marinum infection has never been reported from the subclass. This study investigated the histopathological characteristics and localization of this mycobacterium through molecular analysis of two captive sharks, a scalloped hammerhead Sphyrna lewini and a Japanese bullhead shark Heterodontus japonicus, exhibited in the same aquarium tank. We detected genital mycobacteriosis caused by M. marinum infection using molecular analyses, including polymerase chain reaction (PCR) and DNA sequencing targeting the 60 kDa heat-shock protein gene (hsp65), and peptide nucleic acid–fluorescence in situ hybridization (PNA-FISH) targeting the 16S rRNA gene. Both sharks showed granulomas in connective tissues of the gonads without central necrosis or surrounding fibrous capsules, which is unlike the typical mycobacterial granulomas seen in teleosts. This study reveals that elasmobranchs can be aquatic hosts of M. marinum. Because M. marinum is a representative waterborne NTM and a potential zoonotic agent, cautious and intensive research is needed to overcome a lack of data on the relationship between NTM and the aquatic environment in association with this subclass of Chondrichthyes.     

Utility of a GFP-expressing Barley yellow mosaic virus for analyzing disease resistance genes

The soil-borne plasmodiophorid Polymyxa graminis is a vector for Barley yellow mosaic virus (BaYMV), which can severely damage barley plants. Although 22 disease resistance genes have been identified, only a few have been used for breeding virus-resistant cultivars. Recently, BaYMV strains capable of overcoming the effects of some of these genes have been detected. In this study, green fluorescent protein (GFP)-expressing BaYMV was constructed and used to examine viral dynamics in inoculated barley plants. Leaf inoculations resulted in higher infection rates than root or crown inoculations. Additionally, inoculations of some resistant cultivars produced infections that were similar to those observed in a field test. The results of this study indicate that the GFP-expressing virus is a useful tool for visualizing virus replication and dynamics, and for understanding resistance mechanisms.     

Genetic differentiation between local populations of Ips typographus in the high Tatra Mountains range

Many studies have addressed the question of dispersal potential of Ips typographus, however the approach of molecular ecology have been used only in a few of them. Moreover, none of them has done it on fine scale of just one mountain range. Molecular analysis using a set of six polymorphic microsatellite markers was applied in this study. Adult beetles of I. typographus collected in 22 localities of the High Tatra Mountains were used for the study of genetic differentiation between rapidly expanded local populations of bark beetles after windstorm calamities in spruce stands. Almost none genetic differentiation among local populations or among larger spatial units has been revealed. Analyses did not indicate isolation of populations by distance or formation of population structure. This suggests high potential of I. typographus to disperse over large distances and even the possibility to overcome high mountains ridges.     

Lessons from Two Case Studies of Implementing Traceability in the Dried Salted Fish Industry

The traceability practices in two companies representative of those in a supply chain—one a producer of wet salted cod, the other of dried salted cod—were systematically investigated by a standard procedure. Neither company retained batch identities for all steps in the production process, and so, they did not have an effective traceability system. Although production of dried salted fish is challenging due to the many transformations taking place during production, the use of the basic technique of applying and recording unique identifiers could have provided a traceability system from delivery of wet fish through to the final dried product. This study focused on the critical points for traceability during production of both wet and dried salted fish and the practical challenges this presented.