Hrvoje Mazija

Epidemiological investigation of Chlamydophila psittaci in pigeons and free-living birds in Croatia.

During 2003, 278 adult pigeons (Columba livia) and 54 birds of 11 other free-living species were caught in the various locations in the City of Zagreb, Croatia. Sera from 182 pigeons were tested for the presence of antibodies against Chlamydophila (C.) psittaci by ELISA test and 174 of them (95.6%) were found positive. Because of the high positivity rate in sera, cloacal swabs of 278 pigeons as well as 54 other species of free-living birds were tested for the presence of C. psittaci antigen. Fourty-four of the 278 pigeons (15.83%) were antigen positive, whereas all 54 of the wild birds were negative. Antigen-positive pigeons were euthanised and examined pathomorphologically and cytologically. Findings of specific antibodies and antigen of C. psittaci confirmed the high rate of infection among urban pigeons in the City of Zagreb, fortunately not among other free-living birds. Although the pigeon serovars of C. psittaci are considered to be of moderate pathogenicity for humans, the identification of 15.8% antigen-positive birds represents a potential source of infection to humans, especially for elderly people and immunodeficient patients, as well as for poultry in the Zagreb city area.

Genetic analysis of Newcastle disease virus strains isolated in Bosnia-Herzegovina, Croatia, Slovenia and Yugoslavia, reveals the presence of only a single genotype, V, between 1979 and 2002.

Newcastle disease (ND) epizootics in some European countries after the World War II were caused by ND virus (NDV) of multiple genotypes (IV-VIIa) occurring sequentially and/or simultaneously. This study was carried out to characterise the genetic composition of NDV strains during the outbreaks in the territory of the former Yugoslavia in order to enhance our understanding of the relationships of past epizootics in Europe. Sixty-eight NDV strains isolated between 1979 and 2002 were analysed by restriction enzyme digestion and partial sequencing of the fusion protein gene. All isolates were placed in genotype V, an exotic type, that was introduced to western Europe in 1970. Residue substitution analysis has allowed the recognition of four genetic variants, Vb1-Vb4, and the tracing of their movements. Vb1, a dominant variant in Bulgaria from the late 1970s, was also wide spread in the former Yugoslavia throughout the period under investigation. Vb2, a variant occurring in the neighbouring countries in the early 1970s could be the founder of the epidemic in Yugoslavia and it was present up to the late 1980s. Variants Vb3 and Vb4 could be found only after 1987. In conclusion, the ND outbreaks in Yugoslavia were part of the epizootic wave due to genotype V viruses that started in western Europe in 1970 and became endemic in the region. Inter-country transmission occurred for all variants, and Vb3 and Vb4 might have evolved during the endemic period.

Isolation and molecular biological investigations of avian poxviruses from chickens, a turkey, and a pigeon in Croatia.

Abstract In the last 3 yr, several outbreaks of avian poxviruses (APVs) have been observed in different parts of Croatia. Four strains of APVs, from chickens, a pigeon, and a turkey, were isolated from cutaneous lesions by inoculation onto the chorioallantoic membranes (CAM) of 12-day-old specific-pathogen-free chicken embryos. The resulting proliferative CAM lesions contained eosinophilic cytoplasmic inclusion bodies. The characteristic viral particles of poxvirus were detected in the infected CAM and also in the infected tissues by transmission electron microscopy. Further identification and differentiation of the four various APVs were carried out by the use of a polymerase chain reaction (PCR) combined with restriction enzyme analysis. Using one primer set, which framed a region within the APV 4b core protein gene, it was possible to detect APV-specific DNA from all four tested isolates. PCR results revealed no recognizable differences in size of amplified fragments between the different APVs from chickens, turkey, and pigeon. Restriction enzyme analysis of PCR products using NlaIII showed the same cleavage pattern for turkey and chicken isolates and a different one for the pigeon isolate. Multiplex PCR for direct detection of APV and reticuloendotheliosis virus (REV) was carried out to determine the possible integration of REV in the genome of isolated APVs. The obtained results revealed that REV was present in chicken and turkey strains of poxviruses, whereas the pigeon isolate was negative. It is not known whether the avipoxvirus vaccine strain used in Croatia is contaminated with REV or if the REV is naturally contaminating Croatian field strains of fowl poxvirus. The latter is indicated by the negative REV finding in the pigeon, which was not vaccinated. The results of the present study indicate the reemergence of fowlpox in Croatia, where infections have not been recorded since 1963 and never confirmed etiologically.

Effects of histamine application and water‐immersion stress on gizzard erosion and fattening of broiler chicks

Three groups of Hybro broiler chickens, of which one group were controls, a second treated intragastrically with histamine, and a third stressed by immersion in water, were used in the experiment. Serum activities of aspartate and alanine aminotransferase and creatine kinase were evaluated. Gizzard erosion length, body weight during fattening, slaughter-house or final body weight, food consumption and antibody titre after vaccination for Newcastle disease were also measured. Stress induced severe gizzard erosion and decreased body weight, especially in male birds, whilst both stressed and histamine-treated groups of chicks had decreased antibody titres that were especially pronounced in the stressed animals. Food consumption and aspartate aminotransferase and creatine kinase activities were increased in the stressed chicks.

Immunogenicity and Safety of Queensland V4 and Ulster 2C Strains of Newcastle Disease Virus Given to Maternally Immune, Newly Hatched Chickens by Nebulization

Abstract Commercial chickens with a high level of maternal antibodies for Newcastle disease were vaccinated when newly hatched with Queensland V4 or Ulster 2C Newcastle disease virus (NDV) strains by nebulization. The exposure time to a fine aerosol of vaccine produced with an ultrasonic nebulizer was 60 sec. The chickens were challenged oculonasally with virulent NDV strain Texas GB in weekly intervals up to the 49th day of life. Although protected for several weeks by maternal antibody, they were sufficiently protected thereafter by active immune response to the vaccines. Vaccinal reactions were not observed. Queensland V4 produced higher titers than Ulster 2C and provided better protection to challenge.

Effects of various application routes of Newcastle disease vaccine on specific antibody titres in ostriches.

Newcastle disease (ND) is one of the most important diseases of poultry and other avian species. The usual mean to control ND is specific immunoprophylaxis. Although chickens are routinely vaccinated against ND, vaccination of ostriches is less well understood. We investigated the effect of vaccination against Newcastle disease via different routes on specific antibody titer in 24 adult ostriches, divided into three experimental and one control group. The vaccine was administered in drinking water to the first, by spraying to the second, and oculo-nasally to the third group. The results have indicated antibody production with titers sufficient for humoral immunity in all experimental groups. The strongest immune response was determined in the group vaccinated by spraying. .

Bee Product Apiderm Royal Gel® in Pressure Injuries and Wound Care - Case Report

Bee products are known for thousands years for its beneficial effect on wound healing process [1,2]. Its effectiveness is partly explained only in recent years explaining the multiple bioactivities involved in healing process. It seems that the most important activity of bee products is acidification that promotes healing of deep wounds [3]. Similar report was given by M phande et al. [4] who reported on positive effects of honey and sugar dressing on wound healing. Part of the bee product activity in healing process could be addressed to its strong antibacterial activity directed even to multiresistant strains of bacteria [5]. Honey was also used in treatment of decubitus ulcers [6]. It is obvious that different bee products can act very positive to the healing process in both mechanical wounds as well as to the pressure wounds [7].

Water Soluble Propolis and Royal Jelly Enhance the Antimicrobial Activity of Honeys and Promote the Growth of Human Macrophage Cell Line

Due to the overuse and misuse of antibiotic, an increase in antibiotic resistance of pathogenic bacteria is evolving. Attention should be focused on natural alternatives to antibiotics, like propolis, royal jelly (RJ) and honeys. They all have strong antibacterial properties due to the active substances they contain. This study investigated the effect of combination of water soluble propolis (WSP) Greit120 or fresh royal jelly (F-RJ) (Mižigoj) and Forest honeys as antibacterial against Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis, Acinetobacter baumanii, Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Streptococcus pyogenes, Streptococcus agalactiae and Candida albicans. These substances are also cell growth promoters for human macrophage (TLT) cell line. WSP Greit120, F-RJ (M) and different Forest honeys were prepared in saline as 10% solutions. The antimicrobial activity was expressed as the minimal inhibitory concentration (MIC) in mg/mL. The growth promotion activity was measured at optical density (OD) 595 nm. The combination of WSP Greit120 with different Forest honeys is better than F-RJ (M) in same combination with different Forest honeys. The best antibacterial/antifungal activity was found with the combination of 10% WSP Greit120 in the Forest honey (1:10) from Italy or Spain. When measuring the growth promoting activity of TLT cell line, the best activity was detected at the combination of 10% WSP Greit120 in the Forest honey from Italy (GI3 = 0.796 ± 0.014 and GI5 = 1.133 ± 0.022). Antimicrobial and growth promoting activities are correlated and WSP-dependent.

Multiplication of HVT FC-126 (Herpesvirus turkey) virus in the kidney cell lines of no avian origin

The presented experiments were aimed to cultivate and multiplicate HVT FC-126 in the PLA (Adult pig kidney) and GL-4 (Gerbil kidney) cell lines. Two different HVT FC-126 vaccine strains were used: Marikal SPF (Veterina d.o.o., Croatia) and Lyomarex (Merial, USA). They were adapted to the PLA and GL-4 cell lines. After adaptation, they were titrated on PLA (TCID50 2^4.23) and GL-4 (TCID50 2^4.96). On both cell lines they show similar CPE (cytophatic effect). The difference between them was detected using Real Time PCR, which was also positive by agarose gel analysis for the virus contained in Lyomarex, but not in the Marikal SPF. It can be concluded that both cell lines are sensitive to HVT FC-126 and the virus can be multiplied in high titers though much lower than in the calf intestinal epithelial cell line (CIEB) cells (TCID50 2^7.93).