Selected pathological, immunohistochemical and ultrastructural changes associated with an infection by Diphyllobothrium dendriticum (Nitzsch, 1824) (Cestoda) plerocercoids in Coregonus lavaretus (L.) (Coregonidae)

The pathological changes induced by an infection of Diphyllobothrium dendriticum (Nitzsch, 1824) plerocercoids in powan, Coregonus lavaretus (L.), from Loch Lomond, Scotland, were assessed using immunohistochemical and ultrastructural techniques. In a sample of 26 powan, the occurrence of encysted plerocercoids of D. dendriticum on the outer surface of the stomach was 38.5% (n = 10) with the number of cysts ranging from 4 to 15 and measuring 4.2 +/- 1.0 mm x 3.4 +/- 0.9 mm (mean +/- SD). Histological examination of intestinal samples also revealed plerocercoids (2-21) encapsulated within a proliferation of mesenteric fibrous tissues of the gastric wall and, occasionally, by the gut lamina propria-submucosa and lamina muscularis. In section, cysts were tri-layered and were formed from a series of concentric whorls of fibroblast and collagen fibre-based connective elements. The extent of necrosis within each muscle layer and the serosa of the stomach differed, notably within the latter that was marked by a chronic inflammatory reaction and fibrosis. Within the cyst and around it, a large number of degranulating mast cell/eosinophilic granule cells were seen, in addition to melano-macrophage centres. Immunohistochemical staining of sections of infected stomach revealed a high density of elements, in close proximity to plerocercoids, staining positive for serotonin, bombesin, substance P and galanin. Uninfected material did not present the same levels of activity. Sections through both infected and uninfected tissue were also tested for elements containing vasoactive intestinal peptide, met-enkephalin, calcitonin gene-related peptide, neuropeptide Y and nitric oxide synthase, but these were absent.     

Intestinal microbiota of white shrimp,Litopenaeus vannamei,fed diets containing Bacillus subtilis E20‐fermented soybean meal (FSBM) or an antimicrobial peptide derived from B. subtilis E20‐FSBM

The purpose of the present study is to profile the microfloral structure in the intestines of the white shrimp, Litopenaeus vannamei, after being fed a control diet, in comparison with the diets containing Bacillus subtilis E20‐fermented soybean meal (FSBM) or an antimicrobial peptide (AMP) isolated from B. subtilis E20‐FSBM (diets designated FSBMD and AMPD, respectively) for 60 days. Metagenomic data showed detection of eight phyla, 20 classes, 40 orders, 68 families and 96 genera. Despite no statistically significant difference, an evaluation of microbial diversity recorded higher species richness, Shannon–Weaver diversity index and evenness in the AMPD and FSBMD groups, compared to the control diet group. Venn diagrams showed that 58 of the operational taxonomic units (OTUs) were shared among the control, FSBMD and AMPD groups, but additional beneficial microbes were only found in the FSBMD and AMPD groups. In addition to the augmentation of beneficial bacteria in the FSBMD and AMPD groups, the abundance of potential pathogens, Vibrio and Flavobacterium, was lower in the gut of shrimp fed the FSBMD and AMPD. The results clearly suggest that the FSBMD and AMPD have the capability to change the microfloral structure of shrimp intestines and could be used for disease prevention in shrimp aquaculture.     

Aquaculture of the sand star,Astropecten indicus Döderlein, 1888, as a step toward the sustainable aquaculture of harlequin shrimp,Hymenocera picta Dana, 1852

Ornamental harlequin shrimp, Hymenocera picta, are commonly maintained on live comet sea stars, Linckia multifora. In Thailand, both species are commonly collected from the wild, raising concerns regarding the sustainability of such practices. As a step toward the sustainable aquaculture of both, methods of their culture and nutrition were investigated. Under captive conditions, harlequin shrimp readily predate, and can be maintained upon the astropectinid sand star Astropecten indicus as an alternative live feed during periods when comet sea stars are unavailable. Wild A. indicus subsequently spawned multiple times shortly after their collection; the eggs (300 μm diameter) were transferred to static aquaria. Following the appearance of planktonic bipinnaria, larval sea stars were fed a 50:50 Chaetoceros calcitrans and Isochrysis galbana diet. Eleven days post-transfer, brachiolaria stages began to settle; stages were then fed concentrated I. galbana. When specimens measured 1–2 mm in diameter, they were transferred onto an in-house formulated Amphiprion diet (>50% protein, >16% lipid). Sea stars were graded regularly; they matured and spawned after 6 months. Populations were maintained for more than three successive generations, demonstrating their ready culture under captive conditions, representing the first harvestable aquaculture of a sea star as a step toward supporting the sustainable culture of a primary, marine species.     

Genome-wide insertional mutagenesis of Arabidopsis thaliana

Over 225,000 independent Agrobacterium transferred DNA (T-DNA) insertion events in the genome of the reference plant Arabidopsis thaliana have been created that represent near saturation of the gene space. The precise locations were determined for more than 88,000 T-DNA insertions, which resulted in the identification of mutations in more than 21,700 of the approximately 29,454 predicted Arabidopsis genes. Genome-wide analysis of the distribution of integration events revealed the existence of a large integration site bias at both the chromosome and gene levels. Insertion mutations were identified in genes that are regulated in response to the plant hormone ethylene.     

Systemic Immunosuppression by Methylprednisolone and Pregnancy Rates in Goats Undergoing the Transfer of Cloned Embryos

The presence of the zona pellucida has been perceived as a requirement for the oviductal transfer of cloned embryos at early stages of development while protecting the embryo from an immune system response. We hypothesized that steroid hormone therapy could reduce a potential cellular immune response after the transfer of zona‐free cloned embryos into the oviduct of recipient female goats. In Experiment 1, seven does were used to study the systemic immunosuppressant effect of the methylprednisolone administration (for 3 days) on blood cell counts. Whole blood was collected prior to treatment with methyprednisolone and then on Days 1, 2, 3, 4, 7, 14, 21 and 28 after the first dose of methylprednisolone for the analysis of haematological parameters. Methylprednisolone treatment significantly reduced circulating white blood cells and neutrophils in comparison with pre‐treatment levels, demonstrating a systemic immunosuppressant effect. In Experiment 2, a group of 58 does were used as recipient females to study the effect of administration of methylprednisolone for 3 days on the establishment of pregnancies after the transfer of zona‐free cloned embryos into the oviducts. No effects on pregnancy rates on Day 30 were observed regarding the distinct treatment groups (control vs. methylprednisolone), the source of oocytes (in vivo‐ vs in vitro‐matured) or the presence or absence of the zona pellucida in embryos. In summary, methylprednisolone was effective at inducing a systemic immunosuppressed state in goats, but the treatment prior to embryo transfer did not affect pregnancy rates. Moreover, pregnancy rates were similar between zona‐free and zona‐intact goat cloned embryos.     

Hypoosmotic Swelling Test in Alpaca (Vicugna pacos) Epididymal Spermatozoa

The present study intended to develop the hypoosmotic swelling (HOS) test in alpaca for its use in epididymal spermatozoa, to evaluate the membrane functional integrity and determine an appropriate hypoosmotic solution and whether the incubation time of 15 or 60 min is sufficient for the execution of the test. Hypoosmotic solutions (HS) with the following concentrations were used: 50, 100, 150, 200 and 275 mOsm/kg of sodium citrate tribasic dihydrate and d ‐fructose. Ten microlitres of epididymal sperm sample was mixed in 150 μL of the respective HS and incubated for 15 or 60 min at 38°C. From the proportion of reacted (swollen) spermatozoa, the 150 mOsm/kg HS was the most sensitive (p < 0.05). The exposure times (15 and 60 min) did not have significant differences (p > 0.05) in the proportion of both strong‐ and total‐coiled sperm tails. In conclusion, 150 mOsm/kg HS and 15 min exposure time are optimal to evaluate the plasma membrane functional integrity through the HOS test in alpaca epididymal spermatozoa.     

Comparison of a novel form of zinc with zinc oxide bolus licensed for prevention of facial eczema,and interaction with copper oxide in sheep

To evaluate a novel elemental zinc bolus compared with a registered positive control zinc oxide bolus and assess serum zinc concentrations following concomitant treatment with a capsule containing copper oxide needles. Forty Romney-cross ewes were randomly allocated in a 2 × 2 factorial design study. On Day 0, 20 ewes received novel boluses containing elemental zinc (Investigational Veterinary Product, IVP) while 20 received a zinc oxide bolus (control; CP). Half the animals in each zinc treatment group (n = 10) were treated with a copper oxide needle capsule [Copasure® - Ewe]. Weekly, from Day −7 to 56, all ewes were assessed for signs of photosensitization, and for 10 ewes from each zinc treatment groups, samples were collected for analysis of serum GGT activity, serum zinc concentrations, faecal zinc concentrations and on Days −7 and 56, liver copper concentrations. Multivariable random-effects models assessed the effects of zinc treatment, copper treatment, treatment interactions and time on all analytes. Regression models examined associations between serum and faecal zinc concentrations and GGT activity. Low spore numbers indicated low Pithomyces chartarum challenge. Serum zinc levels were significantly higher in the IVP than in the CP group [p < 0.0001] and varied by time [p < 0.001] and positively associated with faecal zinc concentration [p < 0.001]. Copper treatment did not affect serum zinc [p = 0.82] or faecal zinc [p = 0.92] concentrations. Liver copper concentrations did not differ between zinc treatment groups on Day −7 [p = 0.6] or Day 56 [p = 0.95]. Only the CP/no copper group had no increase in liver copper concentrations.     

Effects of long duration, low dose bronopol exposure on the control of Ichthyophthirius multifiliis (Ciliophora), parasitising rainbow trout (Oncorhynchus mykiss Walbaum)

Ichthyophthirius multifiliis Fouquet, 1876 infections on intensively reared fish stocks can increase rapidly, which if left unmanaged, can result in the heavy loss of stock. The present study explores the efficacy of long duration, low dose (1, 2 and 5 mg L(-1)) treatments of bronopol (marketed as Pyceze?, Novartis Ltd.) in reducing the number of trophonts establishing on juvenile Oncorhynchus mykiss held under small scale culture conditions. The effect of bronopol on the colonisation success of infective theronts was also investigated by adding 2 mg L(-1) bronopol to the water prior and during the infection process. The number of parasites surviving on fish treated this way was compared to groups of fish that only received treatment after infection had occurred. The effect of bronopol on exiting trophonts throughout their external development to the point of theront release was also assessed through the delivery of 1 mg L(-1), 2 mg L(-1) and 5 mg L(-1) bronopol for up to 27 days consecutively (days 9-36 post-infection). The trial showed that a nominal dose of 2 mg L(-1) bronopol administered prior to infection significantly reduced the number of theronts surviving in the water column at the time of the initial challenge by 35-40% (P<0.05). Similarly, doses of 2 and 5 mg L(-1) bronopol administered as the first wave of mature I. multifiliis trophonts exited fish (i.e. day 11 onwards) to develop externally, reduced the number of trophonts establishing on fish as the second cycle of infection by 52-83%. Continuous application of 2 and 5 mg L(-1) bronopol throughout the second and third cycles of I. multifiliis infection gave further reductions of between 90 and 98%. The number of trophonts on the fish in the control tanks and those treated with 1 mg L(-1) and the 2 mg L(-1) dose at the time of initial infection, by comparison, were observed to increase with successive cycles of infection. From these small scale tank trials, this study demonstrates that the strategic, long duration, low dose delivery of drugs like bronopol can significantly reduce the number of trophonts establishing on fish suggesting the potential of this drug at managing I. multifiliis infections.