DISCUSSION

The findings of this retrospective study underscore the critical importance of serial P4 monitoring during the second trimester of canine pregnancy. Our data demonstrate that hypoluteodism is not a stochastic event but rather emerges consistently within a critical window between weeks 5 and 6 of gestation.

Temporal pattern of luteal insufficiency and the critical window

In this cohort, 79.3% of pregnancies required exogenous P4 supplementation during weeks 5–6. This observation corresponds to the well-characterized physiological transition of the canine CL from LH independence to combined LH and prolactin dependence during mid-gestation [18, 19]. Recent large-scale evaluations of P4 dynamics in pregnant bitches similarly reported that, although a gradual decline in serum P4 is expected, abrupt reductions below 5–10 ng/mL during this stage are indicative of luteal insufficiency and are strongly associated with pregnancy loss [20]. Complementary findings by Hinderer _et al. _[4] suggest that, although some individuals may sustain pregnancy at lower concentrations, values below these thresholds, particularly in genetically predisposed breeds, necessitate prompt intervention to prevent embryonic resorption.

The high incidence of luteal failure observed in the SS population during this period reinforces the possibility of breed-specific susceptibility to premature luteolysis or impaired prolactin-mediated luteotrophic signaling. To our knowledge, this study is among the first to quantify a narrow and predictable period of vulnerability (weeks 5–6) in SS. Although a gradual decline in P4 is expected across breeds, the clustering of 79.3% of cases within this interval highlights a breed-specific susceptibility during the transition from LH independence to hormonal dependence. From a clinical perspective, this finding supports targeted hormonal surveillance beginning at days 28–30 post-ovulation in predisposed breeds.

Justification of the 10 ng/mL threshold

The diagnostic threshold of <10 ng/mL used in the present study is more conservative than the 2–5 ng/mL range commonly cited for pregnancy maintenance. However, this threshold was selected because the study population consisted predominantly of high-risk SS with a history of previous pregnancy loss. By intervening immediately after detecting a rapid decline in P4, the objective was to prevent concentrations from reaching levels associated with irreversible placental separation or embryonic resorption.

Although this approach may have resulted in treatment of some bitches that could have maintained pregnancy at lower P4 concentrations, it provided an additional safety margin for breeds predisposed to abrupt luteal failure. Therefore, the threshold adopted in this study should be interpreted as a proactive clinical strategy rather than a strict physiological cutoff.

Efficacy of MPA supplementation

The MPA protocol achieved an 86.2% success rate in maintaining pregnancies to term, which is consistent with previous reports demonstrating the effectiveness of progestin supplementation in managing luteal insufficiency [20]. Administration of MPA effectively compensated for inadequate endogenous P4 production, thereby suppressing premature myometrial activity and maintaining endometrial quiescence.

One of the principal concerns associated with synthetic progestins is the risk of prolonged gestation and fetal compromise. Nevertheless, findings from the present cohort indicate that withdrawal of MPA 24–48 h before the scheduled aesarean section was both safe and effective. This interval appeared sufficient to allow clearance of the exogenous progestin and restoration of parturition mechanisms while preserving fetal viability, consistent with current clinical recommendations [6].

Furthermore, Zhelavskyi et al. [14] emphasized that maintaining physiologically adequate P4 concentrations is the primary determinant of implantation success and fetal survival in bitches with luteal insufficiency. These findings further support the clinical rationale for MPA supplementation in pregnancies at risk of luteal failure.

Clinical rationale for low-dose MPA and the treatment threshold

The absence of virilization among the 47 F puppies contrasts with historical concerns regarding synthetic progestins. This favorable outcome may be attributed to the low dosage used (0.1 mg/kg) and the planned withdrawal of treatment 24–48 h before aesarean section. In addition, elective surgery performed according to fetal maturity, rather than waiting for spontaneous labor, likely minimized the risk of MPA-induced dystocia or prolonged gestation.

However, safety conclusions should be interpreted with caution due to the retrospective design and the absence of an untreated control group. Therefore, the present protocol is more appropriately described as clinically effective under intensive monitoring rather than universally safe.

Although micronized P4 and altrenogest are often preferred because of their shorter half-lives, MPA remains a potent and economical alternative. Importantly, the risk of fetal masculinization associated with synthetic progestins is dose dependent. The use of a low-dose regimen and avoidance of exposure during early organogenesis before day 30 probably contributed to the absence of developmental abnormalities observed in this study. Moreover, the conservative threshold of 10 ng/mL provided a practical clinical buffer for breeds such as SS, in which luteal collapse may occur more abruptly than in mixed-breed populations.

Refractory hypoluteodism and recurrent pregnancy loss

Despite the high overall success rate, the recurrent reproductive failure observed in subject D5 highlights the multifactorial nature of canine pregnancy loss. This bitch experienced pregnancy loss during three consecutive gestations, initially through abortion and subsequently through fetal mummification, despite receiving the same therapeutic protocol that was successful in other animals. These findings suggest refractory hypoluteodism or the presence of concurrent pathological conditions that adversely affected pregnancy maintenance.

Mantziaras and Zakosek Pipan [6] emphasized that pregnancy loss in the bitch is often multifactorial and extends beyond endocrine dysfunction to include infectious agents such as _B. _canis, E. coli, and Salmonella spp., as well as uterine disorders including cystic endometrial hyperplasia and chromosomal abnormalities. Similarly, the occurrence of fetal mummification during the third pregnancy of subject D5 suggests an unfavorable uterine environment or placental insufficiency rather than isolated luteal dysfunction [8, 21].

Therefore, in cases of recurrent reproductive failure despite adequate P4 supplementation, comprehensive diagnostic evaluation for infectious diseases and uterine pathology is strongly recommended [6].

Limitations of the study

An important point of discussion concerns the use of a treatment threshold of <10 ng/mL, which exceeds the traditionally accepted range of 2–5 ng/mL required to maintain pregnancy. This proactive threshold was based not only on the absolute P4 concentration but also on the rate of decline. A rapid decrease from >15 ng/mL to <10 ng/mL over a short period was interpreted as evidence of impending luteal failure. Given the high-risk nature of the cohort, particularly the predominance of SS, early supplementation was initiated to prevent P4 concentrations from reaching levels associated with embryonic resorption.

It is possible that some bitches could have maintained pregnancy at concentrations between 5 and 10 ng/mL without intervention. Consequently, the reported success rate should be interpreted in the context of this preemptive therapeutic strategy. Because the study was retrospective and uncontrolled, the true efficacy of the protocol cannot be conclusively determined.

In clinical practice, withholding treatment from bitches suspected of hypoluteodism presents ethical challenges. Therefore, the study was necessarily observational and descriptive rather than experimentally controlled.

Another limitation was the marked predominance of SS in the study population (21/25 animals). This breed may have an inherent predisposition to luteal insufficiency or exhibit heightened sensitivity during the mid-gestational transition, potentially influencing the observed hormonal patterns. Consequently, the present findings, particularly those concerning the timing and rate of P4 decline, may be most representative of SS, and extrapolation to other breeds should be undertaken cautiously.

The lack of a nontreated control group remains a major limitation. However, considering the history of previous pregnancy loss in a substantial proportion of the cohort, withholding treatment was considered ethically unacceptable by both owners and the institutional ethics committee. Furthermore, the skewed breed distribution indicates that, although the findings are highly relevant to SS, caution is warranted when extending the identified week 5–6 critical window to giant breeds or breeds with substantially different metabolic characteristics.

Clinical implications of the MPA withdrawal protocol

A notable contribution of the present study is the demonstration that short-term withdrawal of MPA before aesarean section (24–48 h) permits endogenous hormonal recovery without compromising fetal viability. Previous reports have highlighted concerns regarding the risk of prolonged gestation associated with MPA administration; however, the standardized withdrawal strategy described herein provides a practical framework for managing elective aesarean section.

These findings support the continued use of MPA as a viable and effective therapeutic option, particularly in resource-limited settings, when combined with intensive monitoring and appropriately timed-withdrawal.

In addition, the present study includes an infographic summarizing the clinical management of canine hypoluteodism (Figure 1). This visual representation outlines the pathophysiology of the condition, the critical gestational window for onset (weeks 5–6), diagnostic criteria, the MPA treatment protocol with planned withdrawal, and the overall reproductive outcomes observed in the cohort.

Figure 1: Infographic illustrating the management of high-risk canine pregnancy as a clinical guide for hypoluteodism.