Immunostimulatory and Antibacterial Effects of Cannabis sativa L. Leaves on Broilers.

The aim of this study was to evaluate the effect of dried Cannabis sativa L. leaves as a phytogenic mixture added to broiler feed on CD4+ and CD8+ T lymphocyte subpopulations, Newcastle disease virus (NDV) antibody titres, and the presence of E. coli in faecal samples. The study was conducted on 100 male Ross 308 broilers, divided into four groups of 25 broilers, for a 42-day research period. The groups were housed separately in boxes on a litter of softwood shavings and were fed starter mixture from day 1 to day 21 and finisher mixture from day 22 to day 42. Industrial hemp (C. sativa) was grown in the Crkvina area, Croatia (latitude: 45°18'46.8″ N; longitude: 15°31'30″ E). The hemp leaves were manually separated, sun-dried, and ground to a powder. The mixture offered to the control group did not contain cannabis leaves, whereas the three experimental groups received mixtures containing mixed cannabis leaves in a quantity of 10 g/kg, 20 g/kg, or 30 g/kg (E_10, E_20, and E_30, respectively). The mean NDV antibody level was uniform in all study groups until post-vaccination day 14 and increased comparably with time. The percentage of CD4+ and CD8+ lymphocytes in the peripheral blood subpopulation showed statistically significant differences (p < 0.001) in the E_20 group as compared with the control group and both the E_10 and E_30 groups throughout the study period. As the broiler age increased, the CD4+-to-CD8+ ratios also increased and were statistically significant (p < 0.0001) on day 42 in all experimental groups as compared to the control group. Comparing the control group with the experimental groups indicated that the bacterial count was lower in broiler groups having received feed with the addition of 20 g/kg and 30 g/kg C. sativa leaves. In conclusion, the C. sativa leaves were found to elicit a favourable immunomodulatory effect on cell-mediated and humoral immune responses in broilers via increased CD4+ and CD8+ lymphocyte subpopulations and higher CD4+:CD8+ cell ratios, thus indicating enhanced immune function capacity. In addition, C. sativa leaves may have complementary effects on the broiler post-vaccination immune response, increase broilers' resistance to infectious diseases, reduce the effect of stress associated with vaccination, and improve broiler health and welfare.

Biogenic amines in selected foods of animal origin obtained from the Croatian retail market.

This study aimed to investigate the presence of eight biogenic amines (BAs): tryptamine (TRP), phenylethylamine (PHE), putrescine (PUT), cadaverine (CAD), histamine (HIS), tyramine (TYR), spermidine (SPD) and spermine (SPM) in cheese, fish & fishery products and meat & meat products obtained from the Croatian retail market. A selective and robust method of high-performance liquid chromatography (HPLC) with diode array detection (DAD) was applied for the determination of BAs in a total of 91 samples in accordance with the performance criteria outlined in the European legislation. A high inter- and intra-food group variability of the amounts of BAs was observed. In the analysed samples, the most represented amines were TYR, HIS, CAD and PUT. Based on the highest content of the most toxic BAs (HIS and TYR) and consequential food safety concerns, the studied food groups can be ranked in the following order: cheese (HIS up to 106.4 mg/kg; TYR up to 206.6 mg/kg), fish &fishery products (HIS up to 98.8 mg/kg; TYR up to 47.9 mg/kg), and meat & meat products (HIS up to 20.0 mg/kg; TYR up to 117.5 mg/kg). The total BA content was significantly higher (p < .05) in fermented in comparison with other food. The study aimed to contribute to the knowledge on BA toxicity and food quality, as well as to support the indispensable future studies of consumption data and exposure assessment, to the end of defining allowable BA concentrations in food.

Demonstration of Usutu virus antibodies in horses, Croatia.

We report the first serological evidence of Usutu virus (USUV) infection in horses in Croatia. During 2011, 1380 horse serum samples from healthy animals were collected from six northern Croatian counties. All samples were first screened for West Nile virus (WNV) immunoglobulin G (IgG) antibodies using an enzyme-linked immunosorbent assay (ELISA). Sixty-nine WNV ELISA-reactive samples were further tested for WNV antibodies by a virus neutralization assay (VN assay) and plaque reduction neutralization test (PRNT), and USUV by a VN assay and tick-borne encephalitis virus (TBEV) antibodies by PRNT. During the same period, 306 human serum samples from patients coming for routine testing with no symptoms of acute febrile disease were tested for USUV IgG using ELISA. Reactive samples were tested for both USUV and WNV using a VN assay. USUV-specific neutralizing antibodies were detected in two of 69 WNV ELISA-reactive horse serum samples. Seropositive animals were found in two different regions of Croatia. One additional sample showed specific WNV-neutralizing antibodies that cross-neutralized USUV. Only one human sample (0.3%) was reactive to USUV antibodies in an ELISA test. In a confirmatory test, WNV-neutralizing antibodies were detected, indicating cross-reactive antibodies with USUV in ELISA. The exposure to USUV was documented in two WNV ELISA-reactive horses at distant locations. These results indicate the presence of USUV in northern Croatia.