Effect of probiotic supplementation and genotype on growth performance, carcass traits, hematological parameters and immunity of growing rabbits under hot environmental conditions

Animals and husbandry

The animals were individually housed in wire cages of a semi-closed rabbitry. All cages were equipped with feeding hoppers and drinking nipples. A total of 80 rabbits aged 8 weeks representing four genetic groups (20 each) of local breed (Jabali, J), imported Spanish breed (V-Line, V) and their crossbreds (¼J¾V and ¾J¼V) were randomly distributed into three dietary probiotic treatments. The animals of the first dietary group (control, 0 g probiotic/t feed) were fed a basal diet without inclusion of probiotics, while the animals of the other two groups were fed diets supplemented with either 200 or 400 g of probiotic/t feed. The utilized probiotic contained 4 × 10⁹ colony-forming units/g of Bacillus subtilis. The commercial basal diet contained 18.5% crude protein, 8.0% crude fiber, 3.0% crude fat and 2250 Kcal metabolizable energy/kg. Feed and water were provided ad libitum.

Growth performance

All rabbits were weighed at the beginning (8 weeks) and the end of the experiment (16 weeks of age). Feed intake was individually recorded throughout the experimental period. Feed conversion ratio (FCR) was calculated as feed consumed divided by weight gain during the experimental period.

Slaughter and carcass characteristics

At the end of the experiment, 56 animals were slaughtered (n = 4–5 animals for each sub-group). Hot carcass, head, skin and viscera were weighed. The carcass was divided into three cuts: fore part, mid part and hind part according to Blasco and Ouhayoun (1993). The results were expressed as percentage of the live body weight. The meat of the hind leg was chemically analyzed for dry matter, crude protein, fat and ash content.

Blood collection and hematological parameters

Two blood samples were collected during slaughter from each rabbit into anticoagulant ethylenediaminetetraacetic acid tubes for determination of hematological parameters and chemical analysis. The hematological parameters were assessed by using Automatic Fully Digital Hematology Analyzer, BC-3000 Plus (Mindray Bio-Medical Electronics Co., Ltd., Shenzhen, China). These parameters included total count of red blood cells (RBC), total count of white blood cells (WBC), hemoglobin (HGB), hematocrit (HCT) and platelets (PLT). The other blood samples were centrifuged for 12 min at 1500 × g and the harvested plasma was stored at –20°C for further analysis. Blood parameters were determined in the plasma using commercial kits (BioMérieux Company, Marcy-l’Étoile, France). Plasma parameters included total protein, albumin and cholesterol.

Assay for Cell-Mediated Immunity

The in vivo response induced by injecting a mitogen was evaluated by injecting phytohemagglutinin P (PHA-P) into the left ear. At 14 weeks of age, 72 rabbits were randomly assigned (six from each sub-group) for this immunocompetence measurement. Each animal was injected intradermally with 100 μg PHA-P (Sigma Chemical Co., St Louis, MO, USA) in 0.1 mL of sterile saline. Upon injection, the site of the needle was marked with permanent black ink to facilitate further measuring. The resultant swelling response in the ear was measured with a constant tension dial micrometer (Ames, Waltham, MA, USA) before injection and at 24, 48 and 72 h after PHA-P injection. The ear swelling was expressed as the difference between the thickness of the ear before and after injection.

Statistical analysis

Data were subjected to a two-way analysis of variance using JMP Ver. 11 (SAS Institute 2013) with probiotic treatment and genetic group as fixed effects. The model was as follows:

where: Y_ijk = the observation taken on the k^th individual; μ = overall mean; G_i = a fixed effect of the i^th genetic group; T_j = a fixed effect of the j^th probiotic treatment; GT_ij = interaction between genetic group and probiotic treatment; e_ijk = random error assumed to be independent and normally distributed with mean = 0 and variance = σ².

All results are presented as mean and the pooled SEM. The significance of difference among the groups was assessed using Duncan's new multiple range test. Significance was set as P < 0.05.

Growth performance

As shown in Table 1, there were no significant differences among dietary probiotic levels or genetic groups for final body weight and weight gain. However, numerical increase in body weight and weight gain was recorded for rabbits fed a diet supplemented with 400 g/t feed compared with the other two groups. Similar results were found by Matusevičius et al. (2006). Shrivastava et al. (2012) reported that the increase in body weight and growth performance in rabbits fed a ration supplemented with mineral mixture and probiotic may be due to increased digestion and absorption through the intestine. Nevertheless, Attia et al. (2012) concluded that the positive effect of probiotics on growth performance resulted from gut microbiota change. In addition, Chandra et al. (2014) attributed the improvement in growth performance to good digestibility and absorption in the ilea. In the present study, the deterioration in body weight and gain of rabbits fed a low level of probiotic (200 g/t feed) is in agreement with the results of Oso et al. (2013). They stated that rabbits fed diets supplemented with probiotics had the lowest weight and gain and they attributed this decrease to instability in the gut integrity. There was a highly significant effect of genetic group and probiotic level on feed intake. V-line consumed significantly (P < 0.01) lower feed compared with the crossbreds. However, a numerical improvement in feed conversion ratio was found in purebreds compared to crossbreds. It is worthy to note that the imported breed (V-line) showed the lowest values of weight gain and feed intake. The results may be due to a negative effect of high temperatures during the experimental period on growth performance for the imported V-line. In this concern, Iraqi et al. (2008) reported that the Egyptian Gabali breed exceeded the V-line breed in their weights and gain. Moreover, the F₁ crossbreds were heavier than the purebreds. Likewise, Al-Dobaib (2010) stated that body weights of crossbred rabbits were heavier than those in V-line purebred.

Carcass characteristics

The results of carcass traits are shown in Table 2. Carcass weight, dressing percentage, cuts of mid part and hind part as a percentage of live body weight were significantly affected by probiotic supplementation. The rabbits given a diet with high levels of probiotics recorded a significantly (P < 0.001) heavier dressing percentage compared with those of the other dietary groups. The same trend was observed for mid part and hind part. Matusevičius et al. (2006) reported that addition of probiotic had an effect on carcass morphology where the treated rabbits showed longer back muscles and higher leg weight. Genetic group had a significant effect (P < 0.01) on carcass traits except for the fore part percentage. The crossbred (¾J¼V) had the heaviest carcass weight, dressing percentage, cuts of fore and mid parts, followed by the local breed (Jabali). It is worthy to note that the V-line breed recorded the lowest dressing percentage among all genetic groups. A similar finding was found by Al-Dobaib et al. (2007), who reported that the carcass weight and dressing percentage had superiority in the crossbreds compared to the V-line.

Data of offal as a percentage of slaughter weight are presented in Table 3. It should be noted that neither probiotic treatment nor genetic group had significant effects. However, liver, heart and kidney are early-maturing organs and rabbits with high gain have an earlier development (Metzger et al. 2006). Ghosh and Mandal (2008) found that there was no significant difference in organ weights among studied breeds.

The chemical composition of meat is shown in Table 4. The probiotic supplementation and genetic group had a significant effect on moisture, dry matter (DM), organic matter (OM) and ash content. Protein and fat % were not significantly affected by probiotic treatment or genetic group. The highest percentages for DM, OM, protein and fat were recorded for rabbits fed a diet containing 400 g/t probiotics, while the lowest figures were found in the control group. Likewise, Chrastinová et al. (2010) reported that natural herb additives and probiotics increased protein and fat contents in rabbit meat. Also, Simonová et al. (2010) reported that meat of rabbits supplemented with oregano and Eleutherococcus had the highest fat content. With respect to genetic group effect, the highest DM, OM, protein and fat contents were found in the meat of the Jabali breed and ¾J¼V crossbred. Generally, meat of crossbred rabbits showed a significant increase (P < 0.05) in ash and OM compared to V-line breed. Similarly, Al-Dobaib et al. (2007) reported that fat content in meat of crossbred rabbits was higher than that of V-line rabbits.

Blood parameters and hematological parameters

The results of blood parameters as affected by probiotic supplementation and genetic group are presented in Table 5. It should be noted that probiotic treatment showed insignificant effects in all traits except for cholesterol. It should be noted that the probiotic supplementation reduced cholesterol content more than it did for the control group. The lowest value was found in rabbits fed a diet containing 400 g probiotic/t feed followed by those fed the low level of probiotics compared to the control. Abdelhady and El-Abasy (2015) found that blood cholesterol was decreased by supplementing with probiotics and they attributed this effect to hydrolase activity of bile salt or to inhibit an enzyme involved in cholesterol synthesis. On the other hand, there is a numerical increase in triglyceride associated with dietary probiotic treatment. A slight increase in total protein level was observed in rabbits fed a diet supplemented with probiotics. The same trend was observed for globulin. It means that the higher level of globulin associated with probiotic treatments may be considered as a good indicator for increases in immunoglobulin and in turn enhances immunity status. Ismail et al. (2002) and Azoz and Al-Kholy (2006) reported that the total protein and globulin levels in blood are considered as good indicators of immunity response and the albumin level reflects liver function. In terms of genetic groups, it could be observed that the crossbred (¼J¾V) had a significantly (P < 0.05) lower globulin level compared with the other genotypes. The Jabali breed recorded a significantly (P < 0.001) lower cholesterol level compared with the other genotypes. Al-Dobaib et al. (2007) found that cholesterol was higher in purebreds than that of crossbreds.

Blood hematological traits are presented in Table 6. It could be concluded that there was no significant difference of probiotic inclusion for all blood parameters studied. However, an increase in platelets was found in rabbits fed a diet containing 400 g probiotic/t feed compared with the other dietary treatments. Also, probiotic supplementation showed higher WBC than in the control group. Attia et al. (2012) found that supplementation of probiotics had a positive effect on increasing HGB, RBC and HCT %. In harmony with the current results, Attia et al. (2015) reported that the inclusion of probiotics in the diet increased the WBC count. Conversely, Atmaca et al. (2014) reported that the decrease in WBC and PLT counts was due to harmful effects on hematopoietic organs. A highly significant (P < 0.02) difference among genetic groups was found for PLT. The highest figure was recorded for Jabali, while the lowest figure was recorded for (¾J¼V) crossbred. Also, the local breed (Jabali) had higher HGB, RBC and HCT figures than the imported breed (V-line); this means that the local breed is more adapted to the hot environmental conditions prevailing in Saudi Arabia. El-Sheikh et al. (2011) reported that genetic group had a significant effect on hematological and biochemical studied traits. They found that native breeds (Black Baladi and Gabali) had higher values than imported breeds (New Zealand and V-Line). They concluded that native breeds had a higher metabolic rate than that of imported breeds.

Cell-mediated immunity

The swelling response due to PHA-P injection in ears of rabbits of different genotypic groups fed various levels of probiotics is shown in Table 7. A statistical (P < 0.05) improvement in cell-mediated immunity after 48 h was recorded in animals fed a diet containing the highest level of probiotics (400 g/t feed) compared to the other treatments. The same trend was observed after 24 h and 72 h but this difference was not significant between dietary probiotic levels. This result is congruent with the finding of El-Kholy et al. (2012), who found that the inclusion of probiotic (Enterococcus faecalis) in the feed of growing New Zealand rabbits greatly enhanced immune response at all studied times. Also, probiotics greatly stimulate cytotoxic and humoral immunity via activation of macrophages and T-cells (Benyacoub et al. 2005; Mountzouris et al. 2007; Parkes 2007). In terms of genetic groups, it should be noted that purebreds significantly (P < 0.05) recorded a higher immune response compared with crossbreds after 24 h of PHA-P injection. This superiority disappeared at later periods. Interaction between probiotic level and genetic group did not exhibit a significant difference at all studied periods.

Conclusion

From the above results, it could be concluded that the level of 400 g probiotic/t feed resulted in slightly increased body weight, weight gain and feed intake. Also, the level 400 g/t feed significantly improved hot carcass weight, dressing percentage and cuts of mid and hind parts. The lowest value of cholesterol and the highest levels of protein, globulin, platelets and WBC were found in rabbits fed a diet supplemented with 400 g probiotic/t feed. It is beneficial to add 400 g probiotic/t feed to improve growth performance, carcass weight, meat quality and immune response in rabbits raised under hot environmental conditions.