Detection of specific antibody responses to vaccination in variable flying foxes (Pteropus hypomelanus)

Megachiropteran bats are biologically important both as endangered species and reservoirs for emerging human pathogens. Reliable detection of antibodies to specific pathogens in bats is thus epidemiologically critical. Eight variable flying foxes (Pteropus hypomelanus) were immunized with 2,4-dinitrophenylated bovine serum albumin (DNP-BSA). Each bat received monthly inoculations for 2 months. Affinity-purified IgG was used for production of polyclonal and monoclonal anti-variable flying fox IgG antibodies. ELISA and western blot analysis were used to monitor immune responses and for assessment of polyclonal and monoclonal antibody species cross-reactivity. Protein G, polyclonal antibodies, and monoclonal antibodies detected specific anti-DNP antibody responses in immunized variable flying foxes, with protein G being the most sensitive, followed by monoclonal antibodies and then polyclonal antibodies. While the polyclonal antibody was found to cross-react well against IgG of all bat species tested, some non-specific background was observed. The monoclonal antibody was found to cross-react well against IgG of six other species in the genus Pteropus and to cross-react less strongly against IgG from Eidolon helvum or Phyllostomus hastatus. Protein G distinguished best between vaccinated and unvaccinated bats, and these results validate the use of protein G for detection of bat IgG. Monoclonal antibodies developed in this study recognized immunoglobulins from other members of the genus Pteropus well, and may be useful in applications where specific detection of Pteropus IgG is needed.     

Consequences of nutritional and husbandry factors on the heat production of rats and broilers. 2. Effect of environmental temperature on the heat production of fully grown broilers and rats

In an experiment with broilers (origin Tetra B) and with rats (albino, Wistar line) with 2 animals each, heat production was ascertained by measuring CO2 production and O2 consumption over 20 minutes after their feeding 18 h and 1 h before the beginning of measuring at ambient temperatures of 30, 25, 20, 15 and 10 degrees C. Every variant was followed through over 6 h/d in 12 measuring sections. The feed amount/ánimal and day was adapted to energy maintenance requirement. At the beginning of the experiments the broilers and rats were 14 and 21 weeks old resp. and weighed 2.2 kg and 220 g resp. The variation of the ambient temperature did not influence the heat production of the broilers. In contrast to this, the time of feeding in relation to the beginning of measuring had a distinct effect on heat production. Whereas a heat production of 342 +/- 34 kJ/kg LW0.75.d was ascertained in the postabsorptive state 18 h after the last feed intake, it increased by 11% to 393 +/- 32 kJ/kg LW0.75.d when measuring began 1 h after feeding. The very act of feed intake increased heat production by 75%. Rats showed a distinct increase of heat production caused by a decreasing ambient temperature. In the temperature range of 30-25 degrees C the increase was shallower than in the range of 25-10 degrees C. Per 1 degrees C below 25 degrees C heat production increased by 30 kJ/kg LW0.75.d. The increase was independent of the metabolism level, which was influenced by the feeding variants. The results are discussed in connection with Rubner's theory of heat compensation.     

Suitability of SSCP-PCR analysis for molecular detection of quinolone resistance in Campylobacter jejuni

Foodborne infections with Campylobacter spp. are increasing, especially antibiotic resistant strains are emerging. Quinolone resistant isolates can cause failure of therapy in severe clinical infections. Molecular characterisation is needed for the detection of resistant variants of C. jejuni. Therefore 23 isolates from poultry and human medicine as well as three control strains were tested for their minimal inhibitory concentration, their Single-Strand-Conformation-Polymorphism (SSCP)-PCR pattern (a method for the detection of resistance determining point mutations), and their sequence of the quinolone resistance determining region (QRDR). Six different SSCP types could be identified: two types for quinolone resistant isolates and other types containing so called silent mutations without influence on the resistance. A genotypic monitoring of the quinolone resistance in C. jejuni can be useful for the early detection of new resistance variants. As a screening method for detection of point mutations in the QRDR the SSCP-PCR can be applied. Compared to other genotypic methods the SSCP-PCR is less time and cost consuming and needs only standard technical equipment.     

Identification of Hammondia heydorni oocysts by a heminested-PCR (hnPCR-AP10) based on the H. heydorni RAPD fragment AP10

Toxoplasma gondii, Hammondia hammondi, Neospora caninum, Neospora hughesi and Hammondia heydorni are members of the Toxoplasmatinae sub-family. They are closely related coccidians with similarly sized oocysts. Molecular diagnostic techniques, especially those based on polymerase chain reaction (PCR), can be successfully applied for the differentiation of Hammondia-like oocysts. In this paper, we describe a rapid and simple method for the identification of H. heydorni oocysts among other members of the Toxoplasmatinae sub-family, using a heminested-PCR (hnPCR-AP10) based on a H. heydorni RAPD fragment available in molecular database. DNA of oocysts of H. heydorni yielded a specific fragment of 289-290 bp in the heminested-PCR assay. No product was yielded when the primers were used for the amplification of DNA extracted from T. gondii, N. caninum, N. hughesi and H. hammondi, thus allowing the differentiation of H. heydorni among other members of the Toxoplasmatinae sub-family. The hnPCR-AP10 was capable of detecting H. heydorni genetic sequences from suspensions with at least 10 oocysts. In conclusion, the hnPCR-AP10 proved to be a reliable method to be used in the identification of H. heydorni oocysts from feces of dogs.     

Comparative growth performance of pigs immunised with a gonadotrophin releasing factor vaccine with surgically castrated pigs and entire boars raised under conventionally managed conditions

The objective of this study was to determine the effect of using a gonadotrophin releasing factor (GnRF) vaccine on growth performance of male pigs. Vaccinated pigs were compared with surgically castrated pigs and entire males under conventionally managed farm conditions. Male pigs were randomly assigned to three treatment groups: pigs surgically castrated during the first week of life (group "C", n=274), pigs immunised twice during the fattening period with a GnRF vaccine, the first when 13-14 weeks of age and the second when 20-21 weeks of age (group "V" n=280) and entire males (group "E", n=56). Average daily weight gain (ADG) was measured during the study. Additionally, feed conversion ratio (FCR) as defined by feed intake/weight gain and feed consumption was assessed during the fattening period. No difference in the ADG during the fattening period could be determined between "C" and "V". From the start of the study until slaughter at study day 160/173 the ADG in group "E" was lower (625 g/day; P) than in group "C" (681 g/day; P < 0.001) and group "V" (675 g/day; P < 0.01). The FCR during the fattening period until study day 160, three days before the first batch of pigs were slaughtered at 24-25 weeks of age, was higher in group"C" (2.72 kg feed intake/kg weight gain) compared with group "V" (2.44 kg feed intake/kg weight gain; P < 0.001) and group "E" (2.44 kg feed intake/kg weight gain; P <0.01). Pigs in group "V" performed similarly to those in group "E" until the second vaccination (day 135) after which time they showed a substantial increase in feed intake and ADG, while retaining an FCR advantage over group C.     

In vitro effects of very low levels of aflatoxin B₁ on free radicals production and bactericidal activity of bovine blood neutrophils

As one of the most potent and hazardous feed/food-originated mycotoxins, aflatoxin (AF) B? is regarded as a potent immunosuppressor in dairy cows. Neutrophils (PMN), as key effector cells against pathogens, have a high potential to kill engulfed microbes. To investigate the in vitro effects of very low doses of AFB? on blood PMN functions, we examined the effects of biologically relevant concentrations of AFB? on the phagocytosis and non-phagocytosis dependent luminol, representative of mainly intracellular free radicals, and isoluminol, representative of mainly extracellular free radicals, chemiluminescence (CL), necrosis and apoptosis of PMN. Isolated blood PMN from healthy dairy cows (n=12) were exposed to 0, 0.01, 0.05 and 0.5 ng/ml of AFB? for 0.5 and 18 h depending on the assay. Further, blood PMN of healthy dairy cows (n=8) were exposed to 0.5 ng/ml of AFB? for 3h and myeloperoxidase (MPO) activity, superoxide anion (O??) production, phagocytosis and killing activities against Staphylococcus (S.) aureus and Escherichia (E.) coli, were examined. Though the effect of extremely low doses of AFB? were less pronounced, at 0.5 ng/ml the production of free radicals was greatly enhanced, especially extracellularly. In contrast to isoluminol CL, the AFB?-treated PMN showed a remarkably impaired phagocytosis-depended luminol CL. PMN necrosis and apoptosis were not affected by AFB?. MPO activity, O?? production, phagocytosis rates and killing of E. coli and S. aureus by AFB?-treated PMN were significantly lower than those of non-treated ones. Our results show the extracellularly pro-oxidant and antiphagocytic properties of very low doses of AFB? for bovine PMN. The scope of the suppressive effects of the in vitro AFB? levels on cellular innate immune functions should be considered for high yielding dairy cows.     

A proposal of clinical breakpoints for amoxicillin applicable to porcine respiratory tract pathogens

In the present position paper, an attempt was made to establish clinical breakpoints of amoxicillin to classify porcine respiratory tract pathogens as susceptible, intermediate or resistant based on their minimum inhibitory concentrations of amoxicillin. For this, a thorough review of the published literature with regard to swine-specific pharmacological data (including dosages of amoxicillin applied and routes of administration used), clinical efficacy, and in vitro susceptibility of the target pathogens was performed. Based on the comparative analysis of the results, the working group "Antibiotic Resistance" of the German Veterinary Medical Society (DVG) proposed to classify porcine respiratory tract pathogens that show MIC values of amoxicillin of < or =0.5microg/ml as "susceptible", those with MICs of 1microg/ml as "intermediate", and those with MICs of > or =2microg/ml as "resistant".     

Leptin, GH, PRL, Insulin and Metabolic Parameters Throughout the Dry Period and Lactation in Dairy Cows

Leptin may play a role in the endocrine-metabolic processes that guarantee the physiological course of lactation in dairy cattle. This study was aimed at determining the changes in plasma concentrations of leptin and some of the main hormones and metabolites involved in the lactogenetic process in high-yielding dairy cows throughout lactation; we also wanted to assess whether leptin secretion is subjected to seasonal influences. Blood samples were collected from 23 Italian Friesian dairy cows from the end of a lactation to the ninth month of the subsequent one; in addition, blood was sampled from 47 dairy cows in different phases of lactation during February and July. Plasma concentrations of leptin, growth hormone (GH), insulin, prolactin (PRL), glucose, non-esterified fatty acids (NEFA) and urea were quantified by either validated radioimmunoassay (RIA) or enzymatic colorimetric methods. At the beginning of lactation, GH concentrations significantly increased, while a significant reduction occurred in leptin and insulin. This endocrine condition, such as the significant increase in NEFA plasma concentrations, is indicative of a marked lipid mobilization. In the more advanced stages of lactation, when both energy and protein balances become positive, leptin plasma concentrations increased, whereas GH and NEFA concentrations declined. During the summer months, a significant increase in leptin plasma concentrations, irrespective of the phase of lactation, was observed. Collectively, our findings suggest that, in dairy cows, leptin may represent a 'metabolic signal' of animal's status of fattening and nutritional level; in addition, leptin seems to be influenced by photoperiod and environmental temperature.