Developmental competence of alpaca oocytes matured in vivo with seminal plasma and following additional in vitro maturation

Silva Huanca, Nancy MilagrosCornelio Díaz, Víctor HugoAuqui Rojas, Luis AntonioHuaman Apaza, Alexis IvanHuanca López, Wilfredo

Abstract The aim of this study was to evaluate the effects of in vivo oocyte maturation using seminal plasma alone or in combination with an additional period of in vitro maturation (IVM) on the developmental competence of alpaca oocytes. The experiment was conducted in Lima, Peru, with twelve adult female alpacas. Follicular ablation of the dominant follicle was performed to initiate a new follicular wave. After 36 hours, a superstimulation protocol with 750 IU of eCG was administered intramuscularly (IM). Four days later, 2 mL of seminal plasma was administered IM to promote in vivo oocyte maturation. Cumulus-oocyte complexes (COCs) were retrieved via ovum pick-up 20 hours post-treatment, morphologically evaluated, and allocated into three groups: (T1) no additional IVM, (T2) 12 hours of additional IVM, and (T3) 18 hours of additional IVM. Oocyte developmental competence was assessed using a 26 µM brilliant cresyl blue (BCB) staining protocol for 90 minutes. COCs were classified as BCB positive (blue ooplasm) or BCB negative (unstained ooplasm) and subsequently denuded and fixed for nuclear maturation assessment via orcein staining. No significant differences (p=0.14) were observed in the percentage of expanded COCs across groups (71.4%, 81.8%, and 54.6% for T1, T2, and T3, respectively). The proportion of COCs reaching the metaphase II (MII) stage was higher (p<0.05) in T3 (54.6%), while the developmental competence rate was greatest (p<0.05) in T2 (100%). However, no differences (p=0.21) were detected in the proportion of BCB positive MII stage COCs across groups. In conclusion, alpaca oocytes matured in vivo with seminal plasma require 12 to 18 hours of IVM to achieve optimal nuclear maturation and developmental competence.