[Determination of the immunoglobulin G supply in the newborn calf]. / Bestimmung der Immunglobulin-G-Versorgung beim neugeborenen Kalb.

A sufficient supply of colostral antibodies within the first hours of life is crucial for the development and the health status in young calves. It is rational to examine the immunoglobulin uptake of single animals, but particularly on a herd basis, during herd controls and consultations. This enables economical calf rearing in accordance with animal welfare. Because of the costly, laboratory-dependent and in part time-consuming direct measurement of the absorbed immunoglobulins using radial immunodiffusion (RID) or ELISA, multiple studies attempted to develop indirect methods, which would be affordable and operational in the field. These aim to draw an inference for the absorbed quantity of colostral antibodies based on other correlated parameters. Multiple validations showed in part significant differences between various methods concerning specificity and sensitivity in comparison to the direct methods. In addition to RID and ELISA, this article presents the measurement of the γ-glutamyltransferase (GGT) activity, the determination of the total serum protein concentration using refractometry and the zinc sulphate turbidity test, and describes the advantages and disadvantages of their application. Refractory measurement and determination of the GGT activity represent a valuable alternative to a laboratory-dependent immunoglobulin G measurement. Nevertheless, there is no ideal rapid test method, such that several influencing factors have to be considered.

[Assessment of immunogenic activity of the cloned human rotavirus A WA strain.]

INTRODUCTION: Rotovirus infection (RVI) caused by the dsRNA-containing virus from genus Rotavirus, Reoviridae family, belonging to group A (RVA), is the cause of severe diarrhea in human and other mammalian species. Vaccination is the most effective way to reduce the incidence of RVI. At present, the effectiveness of using gnotobiotic piglets as a universal model for reproducing human rotavirus infection and assessing the quality of RVI vaccine preparations has been experimentally proven. OBJECTIVES: Evaluation of immunogenic activity of the cloned RVA Wa strain in the new-born Vietnamese potbellied piglets trial. MATERIAL AND METHODS: Development of viral preparations of the cloned human Wa strain PBA, development of human RVA rVP6, ELISA, polymerase chain reaction with reverse transcription, immunization and experimental infection of newborn piglets. RESULTS: The article presents the results of the experiment on double immunization of newborn piglets with native virus preparations with the infection activity 5.5 lg TCID50/ml, 3 cm3 per dose, HRV with adjuvant 500 µg per dose and mock preparation (control group) followed with experimental inoculation of all animals with virulent virus strain Wa G1P[8] human RVA with infectious activity of 5.5 lg TCID50/ml in 5 cm3 dose. Development of clinical signs of disease and animal death were observed only in control group. RT-PCR system to detect RVA RNA in rectal swabs, samples of small intestine and peripheral lymph nodes was developed. ELISA based on obtained human RVA rVP6 was developed and results on RVA-specific IgG-antibodies in serum samples of experimental piglets are presented. CONCLUSION: In the course of the research, a high immunogenic activity of the native and purified virus of the cloned Wa RVA strain Wa was established and the possibility of its use as the main component of the RVI vaccine was confirmed. The possibility of using conventional newborn pigs instead of gnotobiotic piglets as an experimental model was demonstrated.

Correlation of serum IgG concentration in foals and refractometry index of the dam's pre- and post-parturient colostrums: an assessment for failure of passive transfer in foals.

The object of this study was to evaluate the usefulness of measuring the differences in the values of the serum total protein (DVSTP) concentration of foals and the refractometry index (DVRI) of the milk of dams before and after nursing of the colostrum for assessing failure of passive transfer (FPT) in foals. Serum samples from 31 foals were collected before the first nursing and other 1 to 6 times between 4 and 24 hr after birth. Paired colostrum and milk samples were collected from 14 of their dams at the same time. Serum samples were analyzed for IgG concentration using a single radial immunodiffusion (SRID) test (98 samples) and total protein concentration using a temperature-compensating refractometer (98 samples). Colostrum and milk samples were analyzed for refractometry index (RI) using a Brix refractometer (71 samples). DVSTP concentration and DVRI were significantly correlated with serum IgG concentration. The negative predictive values (NPVs) of DVSTP concentration for detecting serum IgG concentrations<400 mg/dl and<800 mg/dl were 98.2% and 91.3% when the cutoff value is set to 0.4 mg/dl and 0.8 mg/dl, respectively. Furthermore, the NPVs of DVRI for detecting serum IgG concentrations<400 mg/dl and<800 mg/dl were 97.3% and 96.3% when the cutoff value is set to 6% and 10%, respectively. The results suggest that measurement of DVRI is useful in assessing FPT as an initial "stall-side" screening test, because it is easy, inexpensive to perform and allows for rapid interpretation.

Behavioral assessment of cognitive function using a translational neonatal piglet model.

Pigs are used in myriad research disciplines related to human health, but no studies have employed the piglet to directly assess cognitive function during the neonatal period. Our objective was to develop a behavioral assay for neonatal piglets to assess learning and memory. At 2-wk of age, piglets were trained to locate a milk reward in an 8-arm radial maze using colored intra-maze cues (acquisition phase, 60-s trials with 8 trials per d for 4d). Cue colors were then reversed and pigs re-tested to assess learning and working memory (reversal phase). Piglets quickly learned the simple associative acquisition task, and proficiency greatly improved throughout reversal testing. To further assess the behavioral assay, piglets received an i.p. injection of saline or polyinosinic:polycytidylic acid (poly I:C; 5mg/kg body weight) immediately preceding reversal testing. Poly I:C-treated piglets exhibited acute sickness behaviors, but observationally, were asymptomatic 12-h post-injection. Pro-inflammatory cytokine mRNA expression was elevated 4-h post-injection in both peripheral and central compartments, and plasma cytokine protein levels were concurrently elevated. Specifically, poly I:C elicited the largest increases in interleukin (IL)-1ß mRNA in the liver, spleen, and hippocampus. At 24-, 48-, and 72-h post-injection (i.e., after acute sickness), poly I:C-treated piglets committed more incorrect arm entries, required more time to complete the reversal task, and moved a greater distance in the maze compared with control piglets. Collectively, these data demonstrate that neonatal piglets are capable of being trained in traditional learning and memory tests, and peripheral immune activation elicits alterations in cognitive processing in the neonate.

Prevalence of failure of passive transfer of immunity in newborn heifer calves and associated management practices on US dairy operations.

Failure of passive transfer of immunity (FPT) in dairy replacement calves has been linked to increased neonatal morbidity and mortality and long-term decreases in productivity. The purpose of this study was to estimate the prevalence of FPT in US dairy heifer calves in 2007 and to use nationally representative data to investigate associations of FPT with colostrum and calf management practices. A cross-sectional study was conducted by the USDA's National Animal Health Monitoring System between January and August 2007. Producers from 394 operations in 17 states completed survey questions about colostrum and calf management practices, and serum samples were collected from 1,816 healthy heifer calves on those operations. Serum immunoglobulin G (IgG) levels were determined by radial immunodiffusion, and calves were classified as having FPT if the IgG concentration was less than 10 mg/mL. To investigate associations between FPT and management practices, a multivariable analysis was completed using a weighted logistic regression model. The estimated prevalence of FPT in US dairy heifer calves was 19.2%. The odds of FPT were higher for calves on operations that pooled colostrum [odds ratio (OR = 2.2)], allowed nursing (OR = 2.4), or hand fed colostrum more than 4 h after birth (OR = 2.7). The odds of FPT were also higher for calves on operations that did not provide a source of heat during cold weather for calves experiencing a dystocia (OR = 1.6), would not seek veterinary assistance when unable to correctly position a calf for delivery (OR = 2.6), or did not routinely monitor serum proteins in calves as a measure of passive transfer (OR = 13.8). The prevalence of FPT in dairy heifer calves has decreased in the last 15 yr, so progress has been made in this important area of calf management. This study identified several management practices associated with FPT that could be targeted for educational campaigns or further research.

Immune response in goats to two commercial foot-and-mouth disease vaccines and the assessment of maternal immunity in their kids.

In this investigation, the immune response of goats to two commercial foot-and-mouth disease vaccines (FMDV) was compared. Highest mean antibody titre was observed on days 60 and 21 in goats vaccinated with two doses of algel (group 1) and oil adjuvant (group 2) quadrivalent vaccines, respectively. There was no significant (P > 0.05) difference in mean antibody titre between the two vaccine groups. However, the antibody titres for type O fell below the protective titres by day 180 and 270 for groups 1 and 2, respectively. The mean maternal antibody titre was 0.610 +/- 0.0 immediately after birth. The highest mean maternal antibody titre was observed at 24 h after birth for all serotypes and then steadily declined. The maternal immunity of kids born to the vaccinated does was persistent up to 90 days after birth. There was no significant (P > 0.05) difference in mean maternal antibody titre between the two groups of goats for all four serotypes throughout the study period. The protective maternal antibody titre for serotype O was maintained only up to 1 week after birth, where for the other three serotypes A, C and Asia1 the protective maternal antibody titre was maintained up to 4 weeks of birth. Oil adjuvant vaccine may be used for control of FMDV in goats and the animals have to be revaccinated after 9 months, whereas the kids must be vaccinated at around 3-4 months after birth. Goats must be included in the FMDV control programmes and the same schedule for cattle can be followed.

Assessment and promotion of judicious antibiotic use on dairy farms in Washington State.

The aims of this study were to describe antibiotic use and biosecurity practices among Washington State dairy producers and to evaluate the effectiveness of a collaborative approach to promoting judicious antibiotic use on dairy farms. In collaboration with a statewide industry group, Washington State dairy producers participated in a written, self-administered survey in 2003. They were then provided several educational interventions followed by a second written survey in 2005. Sixty-five percent (381) of dairy producers completed the 2003 survey. The most commonly cited drugs used for disease treatment were penicillin, ceftiofur, and oxytetracycline. Participants also indicated significant preventive uses with 28% using medicated milk replacer. Most producers appeared to consider intramammary infusion at dry-off to be a treatment rather than a preventative practice. Twenty-three percent of initial respondents indicated at least one extra-label use of antibiotics, yet only half routinely consulted with a veterinarian when doing so. Most agreed that using written protocols for disease treatment could reduce errors, but less than one-third had protocols. After the educational intervention there was a tendency toward reduced use of antibiotic medicated milk replacer: 51% of producers who originally reported using medicated milk replacer discontinued this practice, whereas 12% of producers began using medicated milk replacer between the 2003 and 2005 surveys. Most reported that the resources and educational materials were useful. Areas where additional work is needed include reducing the use of medicated milk replacer, increasing veterinary involvement in antibiotic use decisions, implementing treatment protocols, enhancing biosecurity, and ensuring optimal cow and calf immunity.

Assessment of colostral transfer and systemic availability of immunoglobulin G in new-born foals using a newly developed enzyme-linked immunosorbent assay (ELISA) system.

To measure the immunoglobulin G (IgG) concentration in colostrum, milk and serum samples, a sandwich enzyme-linked immunosorbent assay (ELISA) detection system was developed. The system provided high reproducibility and sensitivity for routine diagnostic purposes. The period of fluctuating serum concentrations of IgG was monitored in new-born foals and their mares for a period of 6 weeks postnatum and postpartum, respectively. All foals received colostrum from their mares. The mean IgG concentration in the precolostral mare serum was approximately 19.0 mg/ml and decreased significantly to 13.8 mg/ml within the first 24 h postpartum. The IgG value fell to a minimum of 11.2 mg/ml by day 21 and increased to 21.6 mg/ml by day 42 postpartum. Within the first 4 h postpartum, mean IgG concentrations of 54.5 mg/ml were measured in the colostrum. A significant decrease to 10.1 mg IgG/ml colostrum was then noted 9-12 h postpartum. The mean IgG concentrations in foal serum increased from 0.3 mg/ml (precolostral value) to 9.6 mg/ml within 5-8 h postnatum. After 13-16 h postnatum, the highest IgG value of 15.7 mg/ml was reached. Over time the mean IgG concentration decreased significantly to 7.9 mg/ml at day 35. At the end of the observation period (day 42 postnatum) the mean IgG concentration once again increased to 11.2 mg/ml serum. In addition, the possible influence of various parameters on IgG concentration were examined. No significant influences could be shown by the breed, mare age, number of pregnancies, days of gestation, month foaled, foal sex, or the different farms. Finally, the cumulative incidence of failure of passive transfer (FPT) defined as IgG levels < 4 mg/ml foal serum, and partial FPT (PFPT) at levels ranging from 4 to 8 mg/ml foal serum was determined. From a total of 70 foals, 10.0% showed FPT and 18.6% showed PFPT.

Assessment of a functional role of auto-anti-idiotypes in idiotype dominance.

We have used a well-defined idiotypic system, the cross-reactive idiotype of A strain (CRIA) (Ab1) idiotype generated in A/J mice injected with arsonate coupled to keyhole limpet hemocyanin (ARS-KLH), to determine the frequency of precursors for auto-anti-idiotypic antibodies (auto-Ab2) in naive and immunized A/J mice by limiting dilution analysis after polyclonal activation by lipopolysaccharide. In naive animals, the precursor frequencies of auto-Ab2 B cells were below the limit of sensitivity of the technique in the majority of A/J mice, and could be detected in only 20% of the animals. Upon immunization with ARS-KLH, a large increase in auto-Ab2 precursor frequency was observed. This shift in frequency was not found when A/J mice were injected with KLH alone, or when BALB/c mice, which do not express the CRIA idiotype, were injected with ARS-KLH. To study the functional role of the auto-Ab2 B cells, we injected neonatal A/J mice with polyclonal rabbit Ab3 antibodies directed against a recurrent idiotype of auto-Ab2. Thereafter, these mice were injected with ARS-KLH. Although the anti-arsonate response level was normal, the CRIA Ab1 expression was reduced tenfold. Thus, the suppression of auto-Ab2 affects Ab1 dominance. We further show that the presence of maternal Ab1 can strongly modify the immune response of the offspring by inducing higher levels of the idiotype after immunization. Furthermore, IgM anti-arsonate antibodies were detected before immunization with antigen. From these data, we conclude that the affinity of antigen alone cannot explain the dominance of CRIA. Network selection is important in the shaping of the available repertoire.

Assembly of recombinant rotavirus proteins into virus-like particles and assessment of vaccine potential.

Rotavirus structural proteins VP4, VP6 and VP7 from Bovine Rotavirus Strain C486 were cloned and expressed in a baculovirus expression system. Combinations of the proteins were assembled into a series of virus-like particles, and a murine model was used to determine the capacity of the recombinant proteins and particles to induce protective immunity. All of the proteins induced humoral immunity as measured by an ELISA against whole virus. However, only the antisera from animals immunized with VP4 neutralized virus and inhibited haemagglutination. Challenge of neonates born to animals immunized with VP4 protein on assembled particles or in cell lysates showed protection against challenge with both homologous (bovine C486) and heterologous (SA-11) strains of rotavirus. In contrast, the offspring of mice immunized with VP6 were only partially protected. Neonates of animals immunized with virus-like particles composed of VP7 assembled on VP6 spherical particles were protected against challenge with the homotypic virus and significantly protected from a heterotypic challenge whereas unassembled VP7 protein provided only partial protection against challenge.

Assessment of neonatal avian inflammatory macrophage function following embryonic cyclophosphamide exposure.

Inflammatory macrophage recruitment and function were examined in 4-5-week-old chickens which had received two doses of cyclophosphamide (CP) or vehicle (dH2O) during late embryogenesis (18 and 19 days of incubation). Mononuclear leukocyte chemotaxis to N-formyl-methionyl-leucyl-phenylalanine (FMLP) and hematological parameters were unchanged in CP-treated vs control chickens. Peritoneal exudate cells (PECs) elicited in response to intraperitoneal (i.p.). Sephadex injection did not differ in CP-treated vs control chickens with respect to cell number, cell type, superoxide anion production, or cell surface expression of Ia and transferrin receptor (TfR) antigens. The CP-treated chickens did exhibit the expected decrease in bursa weight; male chickens exposed to CP also had inhibited testes growth. Although embryonic exposure to CP at this dose results in irreversible bursal damage and subsequent impaired humoral immunity, it appears that there are no long-lasting effects on avian inflammatory macrophage function.

The raising of equine colostrum-deprived foals; maintenance and assessment of specific pathogen (EHV-1/4) free status.

Over a period of two years, a total of 22 full term foals from Welsh Mountain pony mares were raised in conditions that were free from infection by Equid herpesvirus (EHV-1/4). Parturition dates were predicted by monitoring colostrum electrolytes, and the mares allowed to foal naturally under supervision or following induction with intravenous oxytocin. Immediately following birth, foals were separated from their dams and transferred to a specially built, positive pressure isolation unit. They were given antibiotic prophylaxis and fed bovine colostrum during the first 24 h, and then mare's milk replacer until weaned. Out of 22 specific pathogen free (SPF) foals one that had not been given antibiotic prophylaxis died of an E. coli septicaemia aged eight days. Two foals developed a streptococcal upper respiratory tract infection, which responded to antibiotic therapy and did not spread to the rest of the herd. A self limiting upper respiratory tract infection was seen in a fourth foal and mild diarrhoea was observed in six foals. Physical development in all SPF foals appeared normal and behavioural patterns resembled those of conventional handreared foals. Newborn foals were held in a separate quarantine area, within the isolation unit, and checked extensively for evidence of EHV-1/4 infection, before being transferred to the main holding unit. Periodic checks were then made for EHV-1/4 over a period ranging from 2 to 4 months; none of the SPF foals showed evidence of infection with EHV-1/4 in terms of clinical disease, virus isolation, sero-conversion or specific lymphocyte transformation.

Assessment of neutrophil migration, phagocytosis and bactericidal capacity in neonatal foals.

Comparison of neutrophil function was made between 8 clinically normal pony foals (3 to 7 days of age), and their dams. Random migration, stimulated migration to zymosan-activated serum, bacterial phagocytosis and bactericidal capacity of neutrophils were determined in vitro. Random migration was greater (P less than 0.01) and stimulated migration was less (P less than 0.01) in foals than in their dams. Bacterial phagocytosis and bactericidal capacity of neutrophils were not different (P greater than 0.05) between foals and mares. Results of this study suggested that neonatal foals have altered neutrophil locomotion, when compared to their dams.

Non-invasive assessment of endocrine differences in day-old chicks (Gallus domesticus) by analysis of the immunoreactive oestrogen excreted in the egg.

Oestrone, oestradiol and an uncharacterized polar fraction of oestrogenic substances were monitored by radioimmunoassay in Day-0 domesticated chickens (Gallus domesticus) after routine 21.5-day artificial incubation. Paired adrenals and gonads, blood serum and in-ovo excrement (allantoic urates plus cloacal expressions) were collected from 9 female and 10 male chicks. All samples were homogenized, hydrolysed and extracted with ether. Component oestrogens were isolated by celite:ethylene glycol column co-chromatography before assay. All egg excretory oestrogen measurements were higher (P less than 0.001) in females than in males. Oestradiol was the major excretory oestrogen in both sexes; oestradiol:oestrone ratios (mean +/- s.e.m.) in females and males were 18.45 +/- 2.42 and 3.03 +/- 0.37, respectively. Females had higher (P less than 0.01) total oestrogens for gonadal and adrenal homogenates and for blood serum samples than did the males; oestradiol was generally the major component.

A sodium sulfite-precipitation tests for assessment of colostral immunoglobulin transfer to calves.

A sodium sulfite-precipitation test utilizing 3 concentrations of sodium sulfite to evaluate the immune status of neonatal calves allowed scoring of serum immunoglobulin concentrations as less than 5 mg/ml, 5 to 15 mg/ml, and greater than 15 mg/ml. Of the 54 estimations for serums that contained greater than 5 mg/ml, 96% were correct; of the 30 estimations for serums that had between 5 and 15 mg/ml, 90% were correct; and of the 42 estimations for for serums with greater than 15 mg/ml, 93% were correct. It was concluded that this test can be used under field conditions to evaluate the immune status of neonatal calves.