Soil nitrogen dynamics after slurry injection in field trials: Evaluation of a soil sampling strategy

Slurry injection below maize seeds is a rather new application technique developed to improve the nitrogen use efficiency of liquid organic manure. To enable the characterization of the spatial and temporal soil mineral nitrogen (SMN) dynamics after slurry injection, the present study aims to develop an appropriate soil sampling strategy. Three consecutive experiments were conducted. The first testing of the soil sampling approach was conducted in an existing field trial where the slurry was injected down to a depth of 12 cm (upper rim) below the soil surface. The soil profile (75 cm wide) centered below the maize row was sampled grid‐like to a depth of 90 cm. Around the injection zone, soil monoliths (SM) were sampled using a purpose‐built soil shovel. Below the SMs and in the interrow space (15 and 30 cm distance to the row) a standardized auger procedure was performed. The second experiment aimed at improving the sampling strategy with a focus on sample homogenization quality and necessary sample sizes per pooled sample. Furthermore, the risk of a carryover of slurry components along the soil core due to drilling an auger through a slurry band was analyzed. In the third experiment this improved sampling strategy was validated. Results from the first testing of the sampling procedure showed that the strategy is suitable, although some problems occurred (especially the high spread in values among the replications causing high coefficients of variation (CV) of mostly 40–60%). The improvement trial revealed that due to the high gradient of SMN concentration in the direct range of the injection zone an intensive homogenization of these samples is required. Suitable sample sizes (twelve auger samples and six soil monolith samples per pooled sample) have to be collected to obtain reliable SMN values. Drilling an auger through a slurry band to sample subjacent soil layers has to be avoided. Following this enhanced sampling strategy, in the final validation trial the spread in values were considerably reduced and resulted in CV values of mostly < 20%. The developed sampling strategy enables the characterization of the spatial and temporal SMN dynamics when slurry has been band‐injected below a maize row. The method can be transferred to other row crops and different slurry injection spacing.     

Susceptibility of different plant species and tomato cultivars to two isolates of <Emphasis Type="Italic">Pepino mosaic virus</Emphasis>

As Pepino mosaic virus has become a pathogen of major importance in worldwide tomato production, information is needed on possible differences between the sensitivity of cultivars towards infection. Furthermore, it is important what hosts other than Solanaceae may be virus reservoirs and are, therefore, threats for tomato cultivation. Two PepMV isolates (PepMV-Sav, E397, a European tomato isolate and PV-0554, a Peruvian pepino isolate) differing in their origin and virulence were used for several experiments to investigate these issues. The response to mechanical inoculation with PepMV was studied using 25 tomato cultivars, seven indicator plant species, and nine other possible horticultural host plants. Symptom development after infection with PepMV was monitored and the virus was detected by DAS-ELISA and IC-RT-PCR. Garlic and broad bean were shown to be additional hosts of PepMV depending on the virus isolate. Nicotiana benthamiana seems to be the most sensitive indicator among all tested indicator plants developing symptoms. Both PepMV isolates infected all tested tomato cultivars. Development of disease symptoms depended on the cultivar and the virus isolate but symptoms were not visible in all cases. None of the cultivars showed tolerance against the two isolates but two responded with a lower susceptibility at an absorbance level of 0.2 (healthy control 0.09). It was observed that some cultivars grown hydroponically showed also lower losses in biomass and yield. Data indicated a correlation between absorbance level in DAS-ELISA and reduction in total tomato growth.     

Possible cryptic stock structure for minke whales in the North Atlantic: Implications for conservation and management

The minke whale is the last of the great whale species to be hunted in significant numbers. Effective management must include an understanding of how genetic diversity is divided and distributed among putative local populations, and as for many migratory species, this is complicated for the minke whale by large-scale seasonal movement among geographic regions. The problem is that the geographic identity of breeding populations is not known, and instead these whales are predictably found and hunted where different breeding stocks may mix on seasonal feeding grounds. Here we use microsatellite DNA and mtDNA markers to investigate minke whale population structure across the species’ range in the North Atlantic. We found no evidence of geographic structure comparing putative populations in recognized management areas, though some limited structure had been indicated in earlier studies. However, using individual genotypes and likelihood assignment methods, we identified two putative cryptic stocks distributed across the North Atlantic in similar proportions in different regions. Some differences in the proportional representation of these populations may explain some of the apparent differentiation between regions detected previously. The implication would be that minke whales range extensively across the North Atlantic seasonally, but segregate to some extent on at least two breeding grounds. This means that established stock boundaries in the North Atlantic, currently used for management, should be re-considered to ensure the effective conservation of genetic diversity.     

A simplified protocol for molecular identification of Eimeria species in field samples

This study aimed to find a fast, sensitive and efficient protocol for molecular identification of chicken Eimeria spp. in field samples. Various methods for each of the three steps of the protocol were evaluated: oocyst wall rupturing methods, DNA extraction methods, and identification of species-specific DNA sequences by PCR. We then compared and evaluated five complete protocols. Three series of oocyst suspensions of known number of oocysts from Eimeria mitis, Eimeria praecox, Eimeria maxima and Eimeria tenella were prepared and ground using glass beads or mini-pestle. DNA was extracted from ruptured oocysts using commercial systems (GeneReleaser, Qiagen Stoolkit and Prepman) or phenol-chloroform DNA extraction, followed by identification of species-specific ITS-1 sequences by optimised single species PCR assays. The Stoolkit and Prepman protocols showed insufficient repeatability, and the former was also expensive and relatively time-consuming. In contrast, both the GeneReleaser protocol and phenol-chloroform protocols were robust and sensitive, detecting less than 0.4 oocysts of each species per PCR. Finally, we evaluated our new protocol on 68 coccidia positive field samples. Our data suggests that rupturing the oocysts by mini-pestle grinding, preparing the DNA with GeneReleaser, followed by optimised single species PCR assays, makes a robust and sensitive procedure for identifying chicken Eimeria species in field samples. Importantly, it also provides minimal hands-on-time in the pre-PCR process, lower contamination risk and no handling of toxic chemicals.     

The effect of metritis and subclinical hypocalcemia on uterine involution in dairy cows evaluated by sonomicrometry

The objective of this study was to examine the effects of metritis and subclinical hypocalcemia on reduction of uterine size in dairy cows using ultrasonography and sonomicrometry. Four piezoelectric crystals were implanted via laparotomy into the myometrium of the pregnant uterine horn of 12 pluriparous Holstein Friesian cows 3 weeks before the calculated calving date. Sonometric measurements were conducted daily from 2 days before parturition (= Day 0) until Day 14 after calving and then every other day until Day 28. Distances between adjacent crystals were expressed in relation to reference values obtained before calving. The diameter of the formerly pregnant uterine horn was measured using transrectal B-Mode sonography starting on Day 10. Cows were retrospectively divided into the following groups: cows without metritis (M–; n = 7), cows with metritis (M+; n = 5), cows with normocalcemia (SH–; Ca > 2.0 mmol/l on Days 1 to 3; n = 5) and cows with subclinical hypocalcemia (SH+; Ca < 2.0 mmol/l in at least one sample between Days 1 and 3; n = 7). Metritis did not affect (P > 0.05) sonometric measurements, but the diameter of the formerly pregnant horn was larger (P ≤ 0.05) between Days 15 and 21 in M+ cows than in M‒ cows. Reduction in uterine length in hypocalcemic cows was delayed (P ≤ 0.05) between Days 8 and 21 compared with normocalcemic cows, but the uterine horn diameter was not related to calcium status. In conclusion, both diseases affected reduction of uterine size until Day 28. Cows with metritis had a larger uterine diameter, possibly attributable to accumulation of lochia, and cows with subclinical hypocalcemia had delayed reduction of uterine length, presumably related to reduction of myometrial contractility.