Molecular evidence for the lack of transmission of the monogenean Sparicotyle chrysophrii (Monogenea, Polyopisthocotylea) and isopod Ceratothoa oestroides (Crustacea, Cymothoidae) between wild bogue (Boops boops) and cage-reared sea bream (Sparus aurata) and sea bass (Dicentrarchus labrax)

In the Mediterranean area, wild fish have often been suggested as either the reservoirs of the causative agents or at least the carriers of the pathogens responsible for disease outbreaks in cultured fish. However, no epidemiological investigations on actual pathogen/disease interactions between farmed and wild fish have been conducted even for the most important fish pathogens. Only sporadic isolations and identifications of various pathogens in wild fish have been done and real associations with the pathological conditions that exist within the farm environment and vice versa have not been established. Monogenean ectoparasite Sparicotyle chrysophrii and isopod Ceratothoa oestroides are commonly found in the Mediterranean cage-reared sea bream and sea bass and in the surrounding wild fish population. Both species were recognized as pathogens that seasonally inflict serious losses in fingerlings and juveniles of sea bream and sea bass, being potentially propagated and exchanged during wild and caged fish interaction. In order to evaluate the degree of pathogens transfer between wild and farmed fish, we investigated genetic population structures of these two important parasitic pathogens inferred by mtDNA cytochrome oxidase I locus. Parasites isolated from wild and farmed fish on the two most productive Adriatic fish farms showed genetic heterogeneity, contradicting widely accepted hypothesis of cross-contamination, at least in case of S. chrysophrii and C. oestroides. As far as we know, this is the first report that in a study of pathogen transfer molecular evidence was employed to asses the genetic population structure of shared parasites.     

Molecular evidence for cosmopolitan distribution of platyhelminth parasites of tunas (Thunnus spp.)

Global distribution of platyhelminth parasites and their host specificities are not well known. Our hypothesis was that platyhelminth parasites of large pelagic fishes are common around the world. We analysed molecular variation in three different taxa of platyhelminth parasites infecting four species of tunas: yellowfin tuna (Thunnus albacares, Scombridae) from Western Australia, southern bluefin tuna (Thunnus maccoyii, Scombridae) from South Australia, Pacific bluefin tuna (Thunnus orientalis, Scombridae) from Pacific Mexico and northern bluefin tuna (T. thynnus, Scombridae) from two localities in the Mediterranean (Spain and Croatia). Comparisons of ITS2 and partial 28S rDNA demonstrated two congeneric species of blood flukes (Digenea: Sanguinicolidae) from multiple hosts and localities: Cardicola forsteri from southern bluefin and northern bluefin tunas, and Cardicola sp. from Pacific bluefin and northern bluefin tunas; and a gill fluke, Hexostoma thynni (Polyopisthocotylea: Hexostomatidae), from yellowfin, southern bluefin and northern bluefin tunas. Partial 28S rDNA indicates that a second type of fluke on the gills, Capsala sp. (Monopisthocotylea: Capsalidae), occurs on both southern bluefin and Pacific bluefin tunas. This appears to be the first report of conspecific platyhelminth parasites of teleosts with a wide‐ranging geographical distribution that has been confirmed through molecular approaches. Given the brevity of the free‐living larval stage of both taxa of flukes on the gills (H. thynni and Capsala sp.), we conclude that the only feasible hypothesis for the cosmopolitan distribution of these flatworms is migrations of host tunas. Host migration also seems likely to be responsible for the widespread occurrence of the two species of blood flukes (Cardicola spp.), although it is also possible that these were translocated recently by the spread of infected intermediate hosts.     

Essential role of Fkbp6 in male fertility and homologous chromosome pairing in meiosis

Meiosis is a critical stage of gametogenesis in which alignment and synapsis of chromosomal pairs occur, allowing for the recombination of maternal and paternal genomes. Here we show that FK506 binding protein (Fkbp6) localizes to meiotic chromosome cores and regions of homologous chromosome synapsis. Targeted inactivation of Fkbp6 in mice results in aspermic males and the absence of normal pachytene spermatocytes. Moreover, we identified the deletion of Fkbp6 exon 8 as the causative mutation in spontaneously male sterile as/as mutant rats. Loss of Fkbp6 results in abnormal pairing and misalignments between homologous chromosomes, nonhomologous partner switches, and autosynapsis of X chromosome cores in meiotic spermatocytes. Fertility and meiosis are normal in Fkbp6 mutant females. Thus, Fkbp6 is a component of the synaptonemal complex essential for sex-specific fertility and for the fidelity of homologous chromosome pairing in meiosis.     

Effects of peroral insulin and glucose on circulating insulin-like growth factor-I, its binding proteins and thyroid hormones in neonatal calves

There is disagreement in the literature about the ability of neonatal calves to absorb perorally administered insulin. This study evaluated the absorption of a bolus of insulin administered alone or with an energy souce and its effects on the circulating insulin-like growth factor system and thyroid hormones in newborn Holstein-Friesian calves. Within 1 h of dosing, mean serum insulin and triiodothyronine (T3) concentrations had increased considerably, whether the insulin was applied alone (n = 4) or together with glucose (n = 4), accompanied by marked hypoglycemia. No significant changes were observed in control calves (n = 4) given the vehicle solution. Increased serum glucose and T3 concentrations with no change in insulinemia occurred in a 4th group of calves given glucose alone. At 32 h of age and after 3 meals of colostrum there were no differences in glycemia, insulinemia, or proteinemia among the 4 groups of calves examined. Mean serum insulin-like growth factor-I (IGF-I) tended to decrease over this period in the control group. The decrease was more pronounced in the insulin-treated group but absent in both groups that received glucose. These differences were associated with equivalent differences in abundance of the 40-45K IGF-binding protein-3 (IGFBP-3); however, lower molecular mass IGFBPs were not affected. The results show that a pharmacological peroral dose of insulin can lead to rapid systemic alterations in the IGF/IGFBP system in neonatal calves that can be modified by simultaneous administration of a small energy supply in the form of glucose.     

Histopathology of Five Species of Didymocystis spp. (Digenea: Didymozoidae) in Cage-reared Atlantic Bluefin Tuna (Thunnus thynnus thynnus)

The first parasitological examinations of Atlantic bluefin tuna Thunnus thynnus thynnus reared in the Adriatic Sea revealed high numbers of different digenean trematodes, belonging to family Didymozoidae. Five species were the most frequently isolated: Coeliodidymocystis, abdominalis, Didymocystis wedli, Koellikerioides internogastricus, K. intestinalis and K. apicalis. Parasites were isolated from gills, gill rakers, intestine, serosa of pyloric caeca, and inner layers of the fundic region of stomach, encysted in pairs in connective tissue capsules originating from host tissue. Gross pathology revealed no pathological changes. Histological alterations comprised tissue reaction resulting in lymphocyte infiltration and formation of a collagenous envelope around the parasite cysts; the intensity of observed reaction depended on the size of the cyst and on the tissue infected. The most notable changes were observed in the intestine under provocation by K. intestinalis, while the most potentially pathogenic didymozoid under fish rearing conditions is D. wedli, which if present in high abundance can induce secondary bacterial infections.     

Transformation of Petroleum Saturated Hydrocarbons during Soil Bioremediation Experiments

This paper presents transformations of saturated hydrocarbons of petroleum type pollutants during ex situ bioremediation of soil on the pilot heap (halde), during a period of 6 months, within the grounds of Petroleum Refinery Pan?evo (Serbia). Samples for analysis were taken in time intervals of 2 weeks (P₁–P₁₂ samples). Organic substance was extracted by Soxhlet’s method and quantified. Isoprenoid aliphatics, in particular pristane and phytane, and polycyclic aliphatics of sterane and triterpane types in saturated hydrocarbon fractions were analysed by GC-MS (SIM method). Significant amounts of n-alkanes have not been detected. The MS-chromatogram revealed only marginal amounts of pristane and phytane in sample P₁. Pristane and phytane occurred in sample P₈, and in even higher quantities in the final sample P_12. The proceeding bioremediation process was accompanied by the decrease of the relative amounts of pentacyclic terpanes of hopane type, compared to tri- and tetracyclic terpanes. In the initial sample P₁ the distribution of steranes and hopanes follows a pattern, which is characteristic for crude oils. However, their identification by SIM method was not possible in samples P₈ and P₁₂ because of the reduced concentration. The observed changes in the alkane fractions’ compositions may be considered as atypical, referring to the fact that during oil biodegradation under natural conditions, decomposition of isoprenoids occurs much easier and faster than decomposition of polycyclic alkanes of tri-, tetra- and pentacyclic terpane, sterane and diasterane types, after the decomposition of n-alkanes has been almost completed.     

Evaluation of Natural Zeolite as a Material for Permeable Reactive Barrier for Remediation of Zinc-Contaminated Groundwater Based on Column Study

The permeable reactive barrier (PRB) filled with natural zeolite plays the role of a reactive treatment zone for remediation of contaminated groundwater. Based on column lab experiments, the volume of remediated solution, the distribution (K_d) and retardation (R_d) coefficients were evaluated, confirming successful removal and retention of zinc from contaminated groundwater. The effect of hydrodynamic dispersion on zinc capturing by zeolite in PRB was evaluated by the hydrodynamic dispersion coefficient (D_L) and retarded hydrodynamic dispersion coefficient (D_LR) using the Brigham method. For different assumed distances of the barrier, the simulation of one-dimensional zinc concentration profile from the point source through the barrier has been modeled by a simple analytical pulse model. The results show that the flow rate has the most significant effect on the concentration profile, peaks, and broadening of curves. The residence contact time (τ) corresponding to higher K_d and R_d as well as lower D_L and D_LR values outcomes the optimal range of 6.2–9.4 min. This interval corresponds to the experimental performance at the bed length of 8 and 12 cm and flow rate in the range of 6.38–9.57 PV/h. The calculated minimum thickness and longevity confirm the successful application of zeolite as a material in PRB for remediation of zinc contaminated groundwater.     

Production of sexed bovine embryos in vitro can be improved by selection of sperm treatment and co-culture system

The study investigated the effects of sperm sorting, capacitation treatment and co-cultivation on sexed bovine in vitro embryo production. The effect of treatment and co-culture on production of embryos of the preferred sex from unsorted sperm was also studied. Sperm from five breeding bulls was used for fertilization of mature oocytes as follows : Experiment 1, sorted and unsorted sperm (bulls A-E) treated only with heparin in standard co-cultures; Experiment 2, sorted sperm (bulls A-E) treated with heparin-PHE (penicillamine, hypotaurine, and epinephrine) or heparin-caffeine in drop co-cultures; and Experiment 3, unsorted sperm (bull E) treated with either heparin-PHE or heparin-caffeine in both standard and drop co-cultures. In all bulls, treatment with heparin resulted in significantly (p < .05) reduced cleavage and blastocyst rates from sorted sperm, as compared with those from unsorted sperm. In bulls A, B, D and E, treatment of sorted sperm with heparin-PHE in drops significantly increased the blastocyst rate (p < .05). In unsorted sperm of bull E, heparin-PHE treatment in drops resulted in the XX/XY sex ratio inverse to that obtained by heparin-caffeine treatment in standard co-cultures (32.3%/67.7% and 66.7%/33.3%, respectively). In conclusion, the treatment of sorted sperm with heparin-PHE in modified drop co-cultures can be recommended for production of in vitro sexed embryos. The use of unsorted sperm for production of embryos of the preferred sex by selected capacitation treatment and co-culture can be the method of choice in bulls with low IVF yields from sorted sperm.     

Host Switch of <Emphasis Type="Italic">Lamellodiscus elegans</Emphasis> (Monogenea: Monopisthocotylea) and <Emphasis Type="Italic">Sparicotyle chrysophrii</Emphasis> (Monogenea: Polyopisthocotylea) between Cage-reared Sparids

Sharpsnout bream (Diplodus puntazzo) has been used in Adriatic aquaculture for less than a decade, but the decreasing trend of rearing this species will probably result in its complete substitution by more exploited sea bream (Sparus aurata). Only two facilities still rear both fish species in neighbouring cages in monoculture. A switch of parasites was observed between sparids during monitoring of the gill monogeneans of farmed fish. In wild fish of the Adriatic Sea, Lamellodiscus elegans (Monogenea: Monopisthocotylea) has previously been reported in annular (Diplodus annularis), and two-banded sea bream (D. vulgaris) and sharpsnout bream (D. puntazzo), and the present study confirmed its presence also in sea bream, in low prevalence and abundance. The exclusively sea bream monogenean Sparicotyle chrysophrii (Monogenea: Polyopisthocotylea) was also isolated from sharpsnout bream, showing prevalence and abundance values even higher than in its resident host. In the occurrence of L. elegans in sea bream, the opportunistic switch resulted in lower abundance and prevalence than in the original host, while in the second case of switching the monogenean S. chrysophrii showed better reproductive capacity on a new host (sharpsnout bream). Both cases point to the possible enlargement of parasite host range.