Variation in colostral immunoglobulin G concentration in fat tailed sheep and evaluation of methods for estimation of colostral immunoglobulin content

In ruminants, colostrum is a vital source of immunoglobulins that provide passive immunity for their offspring during the neonatal period. It is suggested that colostral immunoglobulin G (IgG) concentration varies between and within breeds and could also be affected by maternal factors. The aim of this study was to investigate possible effects of litter type and ewe parturition number on colostral IgG concentration in two Iranian fat-tailed breeds of sheep (Shaul and Lori Bakhtyari) as well as usefulness of different methods for estimation of IgG concentrations in colostrum. The colostral IgG concentrations were measured in 38 Shaul and 59 Lori Bakhtyari ewes by single radial immunodiffusion, zinc sulphate turbidity and Biuret methods. Measurement of IgG by single radial immunodiffusion revealed that Lori Bakhtyari ewes had significantly (P < 0.05) lower colostral IgG levels (48.82 ± 2.10 mg/ml) than Shaul ewes (62.86 ± 2.48). With regard to the effect of litter type and parturition number, a significant (P < 0.05) difference in IgG concentration of colostrum was only observed between the first (65.17 ± 5.74 mg/ml) and third parturition (41.10 ± 4.60 mg/ml) of Lori Bakhtyari ewes. The colostral IgG concentration was not associated with ewe serum IgG concentration (P > 0.05). The mortality rate was higher in lambs born to ewes with lower IgG in their colostrum. Single radial immunodiffusion did not correlate either with zinc sulphate turbidity method (r = -0.253, P > 0.05) or with Biuret method (r = -0.005, P > 0.05). We can conclude that concentration of colostral IgG could be influenced by breed but not by litter type and parturition number. Colostrum, parturition, litter, radial immunodiffusion, zinc sulphate turbidity, Biuret method Adequate transfer of colostral immunoglobulin (Ig) to newborn lambs is essential as lambs with insufficient amount of antibody are at higher risk of contracting diseases and thus higher mortality rate (Sawyer et al. 1977; Ahmad et al. 2000). The quantity of colostral immunoglobulin transfer could be influenced at different levels including production, ingestion and absorption of IgG. Low production of colostral IgG is one of the major problems culminating in failure of passive immunity transfer (Paulík et al. 1984; Tizard 2004). Influenced by maternal genetics, colostral IgG concentration varies widely between and within breeds (Halliday 1978; Gilbert et al. 1988; Tyler et al. 1999; Gulliksen et al. 2008; Zhang et al. 2009). Concentration of colostral Ig could also be modulated by other factors including parturition number and litter type (Gilbert et al. 1988; Higaki et al. 2012). Single radial immunodiffusion (SRID) has been long applied as an accurate method to measure IgG concentration of colostrum and serum but it is time-consuming and expensive. Other methods have also been considered to estimate serum IgG concentration such as precipitation of globulins by zinc sulphate or assessment of total protein by Biuret method ACTA VET. BRNO 2013, 82: 271–275; doi:10.2754/avb201382030271 Address for correspondence: Gholamreza Nikbakht Brujeni Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran Azadi Avenue, Tehran, Iran Phone: (98) (21) 61117057 Fax: (98) (21) 66427517 E-mail: [email protected] http://actavet.vfu.cz/ (Tizard 2004; Sedlinská et al. 2005; Massimini et al. 2006); however, the usefulness of these test, in comparison to SRID, for prediction of IgG concentration in colostrum remains to be determined. The present study was conducted to determine colostral IgG concentration in Shaul and Lori Bakhtiari breeds, which are two native Iranian fat-tailed sheep (Tavakkolian 2000), and to evaluate the impact of maternal colostral IgG concentration on lamb mortalities. We also investigated the possible effects of litter type and ewe parturition number on colostral IgG concentration as well as correlation of ewe serum IgG concentration with colostral IgG concentration. Finally, the efficacy of zinc sulphate turbidity assay (ZST) and Biuret methods in measurement of colostral IgG concentration in comparison to SRID was assessed. Materials and Methods Animals A total of 38 Shaul and 59 Lori Bakhtyari ewes were randomly selected for the study. Shaul and Lori Bakhtyari ewes were housed at animal research farms of the Faculty of Veterinary Medicine, University of Tehran and Research Centre of Animal Husbandry of Shahrekord (Chaharmahal and Bakhtyari Province, Iran), respectively. The ewes had no apparent signs of illness and were clinically healthy. The body condition of ewes was almost similar across different groups and the average body condition score was similar. The animals were managed in a semi open shed confinement system. Their nutrition consisted of sufficient amounts of alfalfa hay, wheat bran, barley grain, corn silage, and wheat and barley stover. The Lori Bakhtyari ewes also had occasional access to local pasture in summer. The animals were divided into different groups according to their parturition number and litter type (Table 1). It should be noted that total number of Shaul ewes in litter type groups is 29 due to missing information of 9 ewes. During the first month after parturition, mortalities in the offspring of Shaul and Lori Bakhtyari ewes were recorded. Colostrum whey and serum preparation Immediately after parturition, 10 ml of colostrum was obtained from each ewe. For colostrum whey preparation, 1.5 ml of colostrum samples was centrifuged for 5 min at 800 g. Then the upper fat layer was removed and 100 μl of rennin solution (1%) was added to the rest of sample. Skimmed colostrum was kept at 37 °C until the clot separated from whey. Finally the clots were precipitated by centrifugation at 700 g for 5 min. The supernatant was harvested and stored at –20 °C until analysis. Three ml of blood was prepared by jugular venipuncture just after parturition. After blood coagulation at room temperature, serum was separated by centrifugation at 700 g for 10 min and stored at –20 °C until analysis. Immunoglobulin and total protein measurement Total immunoglobulin measurement was performed by a qualitative spectrophotometric zinc sulphate turbidity assay (Pfeiffer et al. 1977). To determine IgG concentration, SRID was carried out in 1% agarose in phosphate 272 Table 1. The colostral IgG concentration (Mean ± SEM) in Shaul and Lori Bakhtyari sheep * Values differ significantly (P < 0.05) †The inconsistency in the total number of Shaul ewes is due to missing information of 9 ewes. Colostral IgG concentration (mg/ml) Shaul breed Lori Bakhtiyari breed Total n mean ± SEM N mean ± SEM n mean ± SEM