Acta Veterinaria Scandinavica 2008, 50:41 http://www.actavetscand.com/content/50/1/41 Table 1: Percentage of sperm morphology of Thai native crossbred (T; T1 – T5) and purebred (F; F1 – Standard-bred; F2 – Warm- blood; F3 and F4 – Thoroughbred) stallions (mean ± SE) Stallion Normal morphology Abnormal head Abnormal midpiece Abnormal tail Proximal cytoplasmic droplet Distal cytoplasmic droplet T1 T2 T3 T4 T5 F1 F2 F3 F4 Mean T (range) Mean F (range) 48.9 ± 1.8 a 43.8 ± 2.4 a,c 60.8 ± 2.7 b 38.3 ± 2.7c 58.3 ± 2.2 b 49.7 ± 1.3 (19 – 75) 40.1 ± 3.6 a 61.5 ± 2.7 b 35.9 ± 3.9 a 58.0 ± 5.1 b 48.1 ± 2.8 (21 – 72) 8.9 ± 0.7 a 14.2 ± 1.5 b 11.5 ± 1.4 a,b 9.2 ± 1.1 a 9.1 ± 1.4 a 10.2 ± 0.5 * (2 – 33) 13.8 ± 1.3 a 10.2 ± 2.1 a 17.3 ± 2.8 b 11.7 ± 1.6 a 13.4 ± 1.1 (4 – 23) 12.7 ± 0.8 a 29.3 ± 1.6 b,c 16.2 ± 1.6 a,c 10.3 ± 0.8 a 19.8 ± 1.6 a 16.5 ± 0.8 * (3 – 41) 31.6 ± 3.4 a,b 14.3 ± 2.5 a 29.9 ± 3.2 b 17.3 ± 2.8 a 23.9 ± 2.1 (9 – 43) 1.1 ± 0.2 a 1.8 ± 0.4 a,b 1.4 ± 0.3 a 2.6 ± 0.5 b,c 1.6 ± 0.3 a,c 1.6 ± 0.1 * (0 – 7) 5.4 ± 1.3 a 2.7 ± 1.0 b 0.3 ± 0.2 c 4.4 ± 1.1 a,b 3.3 ± 0.6 (0 – 13) Significant differences (P < 0.05) within T or F are indicated by different letters (a, b, c, d). Significant differences (P < 0.05) between T and F are indicated by * in superscript 10.9 ± 0.8 a 8.2 ± 1.1 a,b 4.9 ± 1.2 b,c 28.9 ± 2.2 d 3.3 ± 0.5 c 11.4 ± 1.0 * (0 – 42) 3.5 ± 0.8 a 5.7 ± 1.2 a,b 9.3 ± 2.0 b 3.9 ± 0.8 a,b 5.5 ± 0.7 (1 – 17) 17.6 ± 1.4 a 2.9 ± 0.5 b 5.2 ± 1.1 b,c 10.6 ± 1.7 d 7.9 ± 1.1 c,d 10.5 ± 0.8 * (0 – 36) 5.5 ± 2.0 5.5 ± 2.1 7.4 ± 1.5 4.7 ± 1.3 5.8 ± 0.9 (0 – 16) and F stallions, respectively, which closely matches the 50% value that is considered a "normal" average for stal- lions [8]. Our finding is consistent with the 43.4% mor- phologically normal sperm with acceptable fertility [6], but lower than the value for fertile stallions (75.5%) reported by Pesch et al. [7]. For morphologically abnor- mal sperm, high numbers of sperm presented with an abnormal midpiece in both T (16.5%) and F stallions (23.9%). A high proportion of sperm with midpiece abnormalities (25.3%) has also been reported by Voss et al. [32]. However, in this study, the stallions nonetheless achieved acceptable pregnancy rates of 62.5 to 91.7% [32]. One reason why the sperm specimens may have had abnormal midpiece morphology may have been due to a response to environmental insults as seen in bull scrotal insulation studies [33,34]. In addition to impaired epidi- dymal function, insults to spermatocytes or spermatids are also known to result in an increase in cytoplasmic droplet concentration in bull sperm samples [33]. Our study found higher percentages of both proximal and dis- tal cytoplasmic droplets in T stallions than in F stallions. However, these types of abnormality may [7,35] or may not [32,36] affect stallion fertility. A greater impact of sperm abnormality on fertility could be caused by an abnormal head, especially a detached acrosome, as well as by a breakdown in the structural integrity of the plasma membrane and other important organelles. The latter could be identified under transmission electron micros- copy [7,37]. All stallions had more than one billion morphologically normal, progressively motile sperm per ejaculate. On this basis, it might be assumed that all the T stallions were fer- tile, and that they were of comparable fertility to the proven-fertile F stallions. However, their actual fertility or pregnancy rate was not tested in this study. The morphometric characters of normal sperm heads were significantly different among individual T or F stal- lions, and between T and F stallions. Differences in sperm head size within breed have been reported in both Warm- blood [14] and Spanish thoroughbred stallions [15]. Sim- ilarly, differences between breeds have been observed in Arabian, Warm-blood, Thoroughbred and Morgan stal- lions [14]. The results of this study confirm that there is significant variation in normal sperm head characteristics both within and between various breeds of stallions, including the Thai native crossbred. In general, sperm in the T group were larger and rounder than those in the F group were. This may render T sperm more sensitive to certain types of extenders that are commonly employed in cooled storage semen [38]. The cooling rate for stallion sperm can affect sperm motility during storage [39,40]. Sperm of different sizes may undergo different cooling rates during a single procedure. Other researchers have also found that the 'smaller' and 'more elongated' the sperm specimen, the better the sperm's cryoresistance [12]. Thus, sperm head size or shape may be an aspect to consider as part of efforts to improve cooled storage and cryopreservation protocols. Compared to previous studies in which the Harris' hema- toxylin technique was also used, almost all the morpho- metric parameters of F sperm heads in this study were higher than those of both sub-fertile stallions of unclassi- fied breeds [9] and Spanish Thoroughbred stallions [28]. Page 6 of 9 (page number not for citation purposes)