Early Prediction of Corpus Luteum Functionality Using an Imaging Software

The present study aimed to assess the applicability of luteal blood flow data acquiredthrough the use of color Doppler ultrasonography and a post-processing analysistool (ImageJ) for predicting pregnancy in buffaloes (Bubalus bubalis). The experimentwas carried out on 59 multiparous Italian Mediterranean buffaloes that underwentsynchronization of estrus and fixed-time artificial insemination (TAI). Corpus luteumfeatures (size: CLS and blood flow: BFA) were taken from Day 5 to 10 after TAI andretrospectively measured with ImageJ. In the same period, blood samples were taken toassess progesterone (P4) concentrations. Pregnancy diagnosis was carried out on Day45 by ultrasound and confirmed on Day 70 post-TAI. Differences in CLS, BFA, and P4concentrations from Day 5 to 10 after TAI measured between groups were analyzed byANOVA repeated measures as were differences within each day of measuring. Buffaloesthat established a pregnancy (n = 29; 55%) had larger CLS (2.2 ± 0.1 vs. 1.9 ± 0.1 cm2;P < 0.01), higher BFA (0.6 ± 0.0 vs. 0.4 ± 0.0 cm2; P < 0.01), and higher P4 blood level(1.8 ± 0.1 vs. 1.4 ± 0.1; P < 0.01) during Day 5–10 as compared to not-pregnantbuffaloes (n = 22). Throughout the entire period, the first feature that changed betweengroups was P4 blood concentration at Day 7 (1.7 ± 0.1 vs. 1.2 ± 0.1; P < 0.05) followedby BFA at Day 8 (0.6 ± 0.0 vs. 0.5 ± 0.0; P < 0.05), respectively, in pregnant andnot-pregnant animals. The ROC analyses indicated that P4 was able to predict pregnancysince Day 5 (P < 0.05) although a more reliable result could be obtained from Day 8(P < 0.01). At Day 10, it was possible to set a cutoff value for every parameter takeninto account. The logistic regression analysis showed that pregnancy was positivelyinfluenced by P4 concentration (odds ratio 534.127; P < 0.01) and BFA (odds ratio744.893; P < 0.01). In conclusion, the use of color Doppler ultrasonography, togetherwith ImageJ, identified different patterns of BFA between pregnant and not-pregnantbuffaloes starting from Day 8 post-TAI.