Accuracy of early pregnancy diagnosis and determining pregnancy loss using different biomarkers and machine learning applications in dairy cattle.

This study aimed to compare the accuracy of IFN-τ stimulated gene abundance (ISGs) in peripheral blood mononuclear cells (PBMCs), CL blood perfusion by Doppler ultrasound (Doppler-US), plasma concentration of P4 on Day 21 and pregnancy-associated glycoproteins (PAGs) test on Day 25 after timed-artificial insemination (TAI) for early pregnancy diagnosis in dairy cows and heifers. Holstein cows (n = 140) and heifers (n = 32) were subjected to a hormonal synchronization protocol and TAI on Day 0. On Day 21 post-TAI, blood samples were collected for PBMC isolation and plasma concentration of P4. The CL blood perfusion was evaluated by Doppler-US. Plasma samples collected on Day 25 were assayed for PAGs. The abundance of ISGs (ISG15 and RSAD2) in PBMCs was determined by RT-qPCR. Pregnancy was confirmed on Days 32 and 60 post-TAI by B-mode ultrasonography. Statistical analyses were performed by ANOVA using the MIXED procedure and GLIMMIX in SAS software. The pregnancy biomarkers were used to categorize the females as having undergone late luteolysis (LL); early embryonic mortality (EEM); late embryonic mortality (LEM); or late pregnancy loss (LPL). The abundance of ISGs, CL blood perfusion by Doppler-US, and concentrations of P4 on Day 21, and PAGs test on Day 25 were significant (P < 0.05) predictors of early pregnancy in dairy cows and heifers. Dairy cows had a greater (P = 0.01) occurrence of LL than heifers, but there was no difference (P > 0.1) for EEM, LEM, and LPL in heifers compared to cows. Cows with postpartum reproductive issues had a greater (P = 0.008) rate of LEM and a lesser (P = 0.01) rate of LPL compared to cows without reproductive issues. In summary, the CL blood perfusion by Doppler-US had the highest accuracy and the least number of false negatives, suggesting it is the best predictor of pregnancy on Day 21 post-TAI. The PAGs test was the most reliable indicator of pregnancy status on Day 25 post-TAI in dairy heifers and cows. The application of machine learning, specifically the MARS algorithm, shows promise in enhancing the accuracy of predicting early pregnancies in cows.

Microwave Dielectric Response of Bovine Milk as Pregnancy Detection Tool in Dairy Cows.

The most reliable methods for pregnancy diagnosis in dairy herds include rectal palpation, ultrasound examination, and evaluation of plasma progesterone concentrations. However, these methods are expensive, labor-intensive, and invasive. Thus, there is a need to develop a practical, non-invasive, cost-effective method that can be implemented on the farm to detect pregnancy. This study suggests employing microwave dielectric spectroscopy (MDS, 0.5-40 GHz) as a method to evaluate reproduction events in dairy cows. The approach involves the integration of MDS data with information on milk solids to detect pregnancy and identify early embryonic loss in dairy cows. To test the ability to predict pregnancy according to these measurements, milk samples were collected from (i) pregnant and non-pregnant randomly selected cows, (ii) weekly from selected cows (n = 12) before insemination until a positive pregnancy test, and (iii) daily from selected cows (n = 10) prior to insemination until a positive pregnancy test. The results indicated that the dielectric strength of Δε and the relaxation time, τ, exhibited reduced variability in the case of a positive pregnancy diagnosis. Using principal component analysis (PCA), a clear distinction between pregnancy and nonpregnancy status was observed, with improved differentiation upon a higher sampling frequency. Additionally, a neural network machine learning technique was employed to develop a prediction algorithm with an accuracy of 73%. These findings demonstrate that MDS can be used to detect changes in milk upon pregnancy. The developed machine learning provides a broad classification that could be further enhanced with additional data.

Telephone follow-up after early medical abortion using Australia's first low sensitivity urine pregnancy test.

BACKGROUND: Follow-up after early medical abortion (EMA) in Australia often entails tracking serum human chorionic gonadotropin levels or performing ultrasonography in-clinic. In other countries, methods of follow-up such as using a low-sensitivity urine pregnancy test (LSUPT), telephone evaluation and a questionnaire have been demonstrated to be safe and acceptable. AIMS: To evaluate the safety and efficacy of telephone follow-up after EMA using an LSUPT and questionnaire. MATERIALS AND METHODS: A prospective observational cohort study of patients undergoing telephone follow-up after EMA using an LSUPT and questionnaire was conducted from March 26 to July 31, 2020. Outcomes of patients who returned to clinic because of a positive LSUPT were evaluated and adverse event rates were calculated. Routinely collected adverse event information was used to compare complication rates during the evaluation period with that prior to introduction of the LSUPT. RESULTS: During the study period, 2223 patients underwent the new protocol. One hundred and ninety-seven patients had a positive LSUPT at their telephone follow-up. One hundred and thirty-two had an incomplete abortion, 11 had a continuing pregnancy, 53 had a complete abortion and one left the clinic before full assessment. CONCLUSIONS: Introduction of telephone follow-up with an at-home LSUPT reduced the number of patients requiring unnecessary clinic appointments, with over 90% of patients completing their follow-up at home. Complication rates during the study period were found to be at least comparable with previously identified organisational adverse events.

Is Preoperative Urine Human Chorionic Gonadotropin (hCG) Testing Necessary for Pediatric Patients Before Oral and Maxillofacial Surgery Procedures With Sedation?

PURPOSE: Human chorionic gonadotropin (hCG) testing is performed prior to surgical procedures to ensure patient and fetal safety. The purpose of this study was to evaluate the utility of routine pregnancy testing prior to elective outpatient oral and maxillofacial surgery procedures being performed with intravenous sedation (IVS). METHODS: A retrospective cohort study was implemented assessing hCG testing in postmenarche females who underwent elective outpatient oral surgery procedures scheduled with IVS at a tertiary care institution. Medical records were used to identify eligible subjects aged 12 to 45 years. The primary predictor variable was age, and the primary outcome variable was urine hCG test result. Age was divided into groups to reflect early adolescence (12 to 14 years), mid-adolescence (15 to 17 years), late adolescence/early adulthood (18 to 24 years) and adulthood (25+ years). Secondary outcome variables included inability to void for hCG testing, change in anesthetic, case cancellation or rescheduling and were measured over a 2 year period. Descriptive statistics were performed. Relative risk (RR) and Cochran-Armitage test for trend were calculated to determine the statistical significance of age on inability to void. RESULTS: The sample consisted of 5,006 females, with a median age (IQR, range) of 18.0 (3.6, 12.0 to 43.6) years. There was one positive urine hCG result providing a preoperative pregnancy rate of 0.02%. Fourteen of 1,195 subjects (1.2%) over a 2 year period were unable to provide a urine hCG sample. There was a statistically significant trend in inability to void as age groups got older (P = .001). Patients aged 12 to 17 years had an increased risk of being unable to void compared to patients 18 years and older (RR: 14.30, 95% CI: 1.88 to 108.99, P = .01). The total cost of testing over the 11 year observation period was $9,019.59. CONCLUSIONS: The risk of surgical cancellations and delayed care due to patients' inability to void preoperatively plus a lack of any positive preoperative urine hCG findings in patients under 18 years of age in this study, call into question the necessity of routine preoperative hCG screening in pediatric patients presenting for IVS for elective outpatient oral and maxillofacial procedures.

Update on medication abortion follow-up options: many sizes to fit all.

PURPOSE OF REVIEW: To assess the efficacy, benefits, and limitations of available and emerging follow-up options for medication abortion. RECENT FINDINGS: Medication abortion follow-up does not have to be a 'one size fits all' protocol. From most to least invasive, follow-up options include facility-based ultrasound, laboratory-based repeat serum beta-human chorionic gonadotropin (hCG) testing, urine hCG testing (high sensitivity, low sensitivity, and multilevel pregnancy tests), self-assessment with symptom evaluation, and no intervention. Provider or facility-dependent follow-up, including ultrasound and serum testing are effective, but have several limitations, including needing to return to a facility and cost. Remote, client-led follow-up options, such as urine pregnancy testing and symptoms evaluation, are well tolerated and effective for ruling out the rare outcome of ongoing pregnancy after medication abortion and have several advantages. Advantages include being inexpensive and flexible. However, it is important to note that low-sensitivity and multilevel pregnancy tests are not available in all settings. In studies evaluating client-led follow-up with urine pregnancy tests, ongoing pregnancies were identified over half the time with symptoms alone. SUMMARY: Guidelines from several professional organizations have aligned with the evidence and no longer recommend routine office-based follow-up. To ensure care is person-centered, providers should offer follow-up options that align with the comfort, logistical ability, and values of the client.

Efficacy of a low-sensitivity urine pregnancy test for identifying ongoing pregnancy after medication abortion at 64 to 70 days of gestation.

OBJECTIVES: We assessed whether a low-sensitivity pregnancy test is effective at identifying ongoing pregnancy after medication abortion at 64 to 70 days of gestation. STUDY DESIGN: From October 2018 to March 2020, we performed a prospective observational study of participants in England and Wales undergoing medication abortion. Participants were scheduled to return to the clinic 14 ± 3 days after mifepristone administration to perform a low-sensitivity pregnancy test (human chorionic gonadotropin threshold of 1000 mIU/mL) and symptom checklist, and state whether they thought the abortion was complete. Clinicians also assessed the low-sensitivity pregnancy test and performed an ultrasound to determine abortion status. We calculated the sensitivity, specificity, negative and positive predictive value of the low-sensitivity pregnancy test (with and without a symptom checklist) for detecting ongoing pregnancy. RESULTS: We enrolled 757 participants. Thirty-one did not progress to abortion and 558 (76.9%) completed follow-up. Most (79.6%) attended per-protocol; 22 (3.9%) attended earlier than 11 days and 92 (16.5%) later than 17 days. Thirteen participants (2.3%) had an ongoing pregnancy. The low-sensitivity pregnancy test correctly identified all the ongoing pregnancies (sensitivity = 100%; specificity = 84.8%; negative predictive value = 100%; positive predictive value = 13.5%). The symptom checklist alone had a sensitivity of 76.9% and a negative predictive value of 99.4% for identifying ongoing pregnancies. Participants and clinicians agreed on the interpretation of the low-sensitivity pregnancy test 94.6% of the time. CONCLUSIONS: Patient self-assessment of a low-sensitivity pregnancy test after medication abortion between 64- and 70 days' gestation has high sensitivity and negative predictive value for identification of ongoing pregnancy. IMPLICATIONS: Patients can be offered a low-sensitivity pregnancy test to assess for ongoing pregnancy after medication abortion up to 70 days of gestation thereby reducing the need for in-person visits. Services should be prepared to provide in-person assessments after positive or inconclusive results to ensure early identification of abortion complications.

Aptamers based sensing of pregnancy associated glycoproteins (PAG) of bovine for early pregnancy detection.

Tosyl activated magnetic beads were used for aptamer selection against PAG- 7 and 18 proteins of bovine origin. PAG proteins were immobilized on beads with further addition of biotin tagged aptamer library. The recognition of aptamers with PAG was identified by ST-HRP based approach which was colorimetric in nature. The selected aptamers were sequenced and at the same time several new aptamers were identified. Later M-fold structure and G-quadruplex score of aptamers were analyzed for their selection. Those aptamers having high G value and complex structure were chosen. In dot blot assay, aptamers recognized PAG protein in an animal after 42 days of artificial insemination which later given birth to a healthy calf. Further the cross reactivity with serum of 0th day animal (post AI) or with non pregnant animal serum was minimal. Aptamers have also shown interaction with PAG protein of buffalo origin. These selected aptamers have commercial application especially in development of biosensors for early detection of pregnancy in bovine.

Prevalence and correlates of pregnancy self-testing among pregnant women attending antenatal care in western Kenya.

In sub-Saharan Africa little is known about how often women use pregnancy self-tests or characteristics of these women despite evidence that pregnancy self-testing is associated with early antenatal care (ANC) initiation. Understanding the characteristics of women who use pregnancy self-tests can facilitate more targeted efforts to improve pregnancy testing experiences and entry into the ANC pathway. We conducted a cross-sectional survey among pregnant women enrolling in a pre-exposure prophylaxis (PrEP) implementation study to determine the prevalence and factors associated with pregnancy self-testing among women in western Kenya. Overall, in our study population, 17% of women obtained a pregnancy self-test from a pharmacy. Pregnancy test use was higher among employed women, women with secondary and college-level educated partners, and women who spent 30 minutes or less traveling to the maternal and child health (MCH) clinic. The most reported reasons for non-use of pregnancy self-tests included not thinking it was necessary, lack of knowledge, and money to pay for the test. Future research should focus on understanding the knowledge and attitudes of women toward pregnancy self-testing as well as developing community-based models to improve access to pregnancy testing and ANC.

Algorithms predicting gestational stage from the maternal steroid metabolome of mares.

Hormone secretion by the maternal ovaries, trophoblast/placenta and fetus occurs sequentially, creating distinct steroid metabolomic 'signatures' in systemic blood of pregnant mares that vary with gestational stage. Algorithms were developed to predict the gestational day (GD) from the maternal steroid metabolome (nine steroids; pregnenolone (P5), progesterone (P4), 5α-dihydroprogesterone (DHP), 17α-hydroxyprogesterone, allopregnanolone, 20α-hydroxy-DHP, 3ß,20α-dihydroxy-DHP, DHEA and androstenedione) determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) of eight thoroughbred mares sampled longitudinally throughout pregnancy. A physiologically based model was developed to infer rates of steroid secretion during chorionic gonadotropin secretion, the luteo-placental shift and by the equine feto-placenta unit, demonstrating more variability in P5 and DHP than P4. The average of four empirical models, using nine steroids to predict GD, was calibrated (five mares, R2 = 0.94, RMSE = 20 days) and validated (three mares, R2 = 0.84, RMSE = 32 days). Validation performance was improved using paired samples taken 14 or 30 days apart (RMSE = 29 and 19 days, respectively). A second validation used an independent dataset (single serum samples from 56 mixed breed mares, RMSE = 79 days) and an additional longitudinal subset from the same population sampled monthly throughout gestation (seven mares, RMSE = 42 days). Again, using paired samples improved model performance (RMSE = 32.5 days). Despite less predictive performance of the mixed breed than the thoroughbred datasets, these models demonstrate the feasibility and potential for using maternal steroid metabolomic algorithms to estimate the stage of gestation in pregnant mares and perhaps monitor fetal development.

Thermal features, ambient temperature and hair coat lengths: Limitations of infrared imaging in pregnant primitive breed mares within a year.

Infrared thermography is a non-invasive technique which allows to distinguish between pregnant and non-pregnant animals. Detecting accurate body surface temperatures can be challenging due to external factors altering thermograph measurements. This study aimed to determine the associations between the ambient temperature, the hair coat features and the temperatures of mares' abdomens. It compared pregnant and non-pregnant mares throughout 11 months. The research was carried out on 40 Konik Polski mares, which were divided into pregnant and non-pregnant groups. The temperature (Tmax, maximal; Taver, average; Tmin, minimal) of the mares' abdomen was evaluated in two regions of interest: the whole area of the lateral surface of the mares' abdomen (Px1) and the flank area of the lateral surface of mares' abdomen (Px2). During the increasing period, the slopes in the linear regression equation did not differ significantly for ambient (Tamb) and surface temperatures in both groups. In the decreasing period, the slopes did not differ significantly for Tamb and Tmax in the non-pregnant group. They also did not differ for Tamb and Taver in Px1 and Tamb and Tmin in Px1 in both pregnant and non-pregnant groups respectively. Other slopes varied significantly (p < .001). There was no evidence of parallel changes in hair coat features and measured temperatures. The flank area appears more suitable for thermal imaging in pregnant mares due to the seasonal fluctuations in hair coat lengths.

EVALUATION OF A VISUAL ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA) FOR PREGNANCY DETECTION IN FOUR UNGULATE SPECIES.

This study evaluated the use of a commercially available, visual enzyme-linked immunosorbent assay (ELISA) for diagnosis of pregnancy in okapi (Okapia johnstoni), gerenuk (Litocranius walleri), eastern giant eland (Tragelaphus derbianus spp. gigas), and dama gazelle (Nanger dama). This assay has been validated for use in domestic cattle, sheep, goats, and water buffalo. Unlike other blood-based pregnancy associated glycoprotein (PAG) detection methods, this assay does not require sophisticated laboratory equipment for detection or interpretation and can therefore be utilized in many settings. Banked serum samples from gerenuk (n = 11), giant eland (n = 4), dama gazelle (n = 33) and okapi (n = 3) were tested, and a pregnant and nonpregnant sample from each individual were included. The ELISA showed 100% sensitivity and specificity in gerenuk and giant eland samples, and 0% sensitivity in dama gazelle and okapi samples. Using this assay, pregnancy was detected by 7-8 wk gestation in gerenuk and 6 wk in giant eland. These results are consistent with previous studies that were able to accurately detect pregnancy in other members of the family Bovidae, but it is possible that PAGs present in okapi and dama gazelle are structurally dissimilar relative to the intended test target, and are therefore unrecognizable using this test. The faint positivity in the dama gazelle assays may be due to cross-reactivity with other proteins in the sample, or due to inconsistent binding with the dama gazelle PAG. This ELISA appears to be an accurate, rapid, and inexpensive method of point-of-care pregnancy diagnosis in gerenuk and giant eland, but not okapi and dama gazelle. Additional studies should be pursued to further characterize the limits of pregnancy detection using this assay in gerenuk and giant eland, and to investigate the validity of this test in other nondomestic ruminant species.

External validation of models to predict the outcome of pregnancies of unknown location: a multicentre cohort study.

OBJECTIVE: To validate externally five approaches to predict ectopic pregnancy (EP) in pregnancies of unknown location (PUL): the M6P and M6NP risk models, the two-step triage strategy (2ST, which incorporates M6P), the M4 risk model, and beta human chorionic gonadotropin ratio cut-offs (BhCG-RC). DESIGN: Secondary analysis of a prospective cohort study. SETTING: Eight UK early pregnancy assessment units. POPULATION: Women presenting with a PUL and BhCG >25 IU/l. METHODS: Women were managed using the 2ST protocol: PUL were classified as low risk of EP if presenting progesterone ≤2 nmol/l; the remaining cases returned 2 days later for triage based on M6P. EP risk ≥5% was used to classify PUL as high risk. Missing values were imputed, and predictions for the five approaches were calculated post hoc. We meta-analysed centre-specific results. MAIN OUTCOME MEASURES: Discrimination, calibration and clinical utility (decision curve analysis) for predicting EP. RESULTS: Of 2899 eligible women, the primary analysis excluded 297 (10%) women who were lost to follow up. The area under the ROC curve for EP was 0.89 (95% CI 0.86-0.91) for M6P, 0.88 (0.86-0.90) for 2ST, 0.86 (0.83-0.88) for M6NP and 0.82 (0.78-0.85) for M4. Sensitivities for EP were 96% (M6P), 94% (2ST), 92% (N6NP), 80% (M4) and 58% (BhCG-RC); false-positive rates were 35%, 33%, 39%, 24% and 13%. M6P and 2ST had the best clinical utility and good overall calibration, with modest variability between centres. CONCLUSIONS: 2ST and M6P performed best for prediction and triage in PUL. TWEETABLE ABSTRACT: The M6 model, as part of a two-step triage strategy, is the best approach to characterise and triage PULs.

Circular RNA Sequencing of Maternal Platelets: A Novel Tool for the Identification of Pregnancy-Specific Biomarkers.

BACKGROUND: In the first trimester of pregnancy, the maternal platelet is directly involved in a positive feedback mechanism that facilitates invasion of the extravillous trophoblast into the maternal spiral arteries. Dysfunctional trophoblast invasion with defective deep placentation is primordial in the etiology of the "great obstetrical syndromes." METHODS: In this proof-of-concept study, using transcriptome analysis of circular RNA (circRNA) following RNA sequencing of maternal platelets, we tested whether pregnancy-specific circRNA markers could be identified in the first trimester of normal pregnancies. Differential transcript expression analysis of circRNAs, as predicted by Accurate CircRNA Finder Suite, CircRNA Identifier (version 2), and Known and Novel Isoform Explorer, was done using thromboSeq.R with variation of multiple settings. Test performance was checked for (a) de novo circRNA identification using the novel platelet-specific Plt-circR4 as a positive control, (b) complete segregation of groups (pregnant vs nonpregnant) after heat map-dendrogram clustering, (c) identification of pregnancy-specific circRNA markers at a false discovery rate (FDR) <0.05, and (d) confirmation of differentially expressed circRNA markers with an FDR <0.05 by an independent method, reverse transcription-quantitative PCR. RESULTS: Of the differentially expressed circRNAs with P values <0.05, 41 circRNAs were upregulated (logFC >2), and 52 circRNAs were downregulated (logFC less than -2) in first-trimester platelet RNA. Of these, nuclear receptor-interacting protein 1 circRNA covering exons 2 and 3 of the 5'-untranslated region was pregnancy specific with upregulation in first-trimester maternal platelets compared to nonpregnant controls. CONCLUSION: CircRNA sequencing of first-trimester maternal platelets permits the identification of novel pregnancy-specific RNA biomarkers. Future use could include the assessment of maternal and fetal well-being.

Efficacy of a commercial test kit to determine early pregnancy in cows using whole blood and blood serum.

The objective of this study was to determine the efficacy of the Fassisi®BoviPreg visual test kit (Fassisi®BoviPreg) in determining early pregnancies in cows by measuring the pregnancy-associated glycoprotein (PAG) in whole blood and blood serum. The study was conducted on 50 cows, the artificial insemination (AI) dates of which were designated as day 0. Pregnancy diagnosis was performed with transrectal ultrasonography (USG), and serum samples were simultaneously collected and used with Fassisi®BoviPreg to determine pregnancies on 30 days after AI. The results of the Fassisi®BoviPreg test on serum and whole blood samples, respectively, on 30 days after AI were as follows: sensitivity, 61.54% and 50.0%; specificity, 79.17% and 75%; accuracy, 70.0% and 62.0%; positive predictive values, 76.2% and 68.4%; negative predictive values, 65.5% and 58.1%; false-positive diagnoses, 23.8% and 31.6%; and false-negative diagnoses, 34.5% and 41.9%. On day 50 after AI, sensitivities were 63.64% and 50.0%, specificities were 100.0% and 100.0%, and accuracies were 75.0% and 65.62% in serum and whole blood, respectively. Higher pregnancy rates were obtained using Fassisi®BoviPreg in cows with ≥ 5 ng/mL P4 (P < 0.001). In conclusion, the results from the Fassisi®BoviPreg tests on cows on 30 and 50 days after AI showed that the use of blood serum is more accurate and suitable than that of whole blood. The results also showed a higher confidence level in specificity 50 days after AI.

External validation of risk prediction model M4 in an Australian population: Rationalising the management of pregnancies of unknown location.

BACKGROUND: The prediction model M4 can successfully classify pregnancy of unknown location (PUL) into a low- or high-risk group in developing ectopic pregnancy. M4 was validated in UK centres but in very few other countries outside UK. AIM: To validate the M4 model's ability to correctly classify PULs in a cohort of Australian women. MATERIALS AND METHODS: A retrospective analysis of women classified with PUL, attending a Sydney-based teaching hospital between 2006 and 2018. The reference standard was the final characterisation of PUL: failed PUL (FPUL) or intrauterine pregnancy (IUP; low risk) vs ectopic pregnancy (EP) or persistent PUL (PPUL; high risk). Each patient was entered into the M4 model calculator and an estimated risk of FPUL/IUP or EP/PPUL was recorded. Diagnostic accuracy of the M4 model was evaluated. RESULTS: Of 9077 consecutive women who underwent transvaginal sonography, 713 (7.9%) classified with a PUL. Six hundred and seventy-seven (95.0%) had complete study data and were included. Final outcomes were: 422 (62.3%) FPULs, 150 (22.2%) IUPs, 105 (15.5%) EPs and PPULs. The M4 model classified 455 (67.2%) as low-risk PULs of which 434 (95.4%) were FPULs/IUPs and 21 (4.6%) were EPs or PPULs. EPs/PPULs were correctly classified with sensitivity of 80.0% (95% CI 71.1-86.5%), specificity of 75.9% (95% CI 72.2-79.3%), positive predictive value of 37.8% (95% CI 33.8-42.1%) and negative predictive value of 95.3% (95% CI 93.1-96.9%). CONCLUSIONS: We have externally validated the prediction model M4. It classified 67.2% of PULs as low risk, of which 95.4% were later characterised as FPULs or IUPs while still classifying 80.0% of EPs as high risk.

Applied use of interferon-tau stimulated genes expression in polymorphonuclear cells to detect pregnancy compared to other early predictors in beef cattle.

We aimed to evaluate the accuracy of Interferon-tau stimulated genes (ISG) abundance in peripheral blood polymorphonuclear cells (PMNs) on D20 after fixed-time artificial insemination (FTAI; D0) as a pregnancy diagnosis method against CL evaluation by Doppler ultrasonography and progesterone (P4) concentrations on D20, as well as Pregnancy Associated Glycoproteins (PAG) concentrations on D25. Additionally, we evaluated the potential of ISG abundance in PMNs as pregnancy loss predictors. Nelore heifers (n = 103) and cows (n = 144) underwent estrous synchronization and were artificially inseminated on D0. Pregnancy was diagnosed by B-mode ultrasonography on D30 and D70, and after the final diagnosis, females were classified in four groups: Pregnant; Non-pregnant; Functional CL on D20 but non-pregnant (CL-NP) and Pregnancy loss between D30 and D70 (PL). After determining cutoff values, the Sensitivity (SE), Specificity (SP), Positive Predictive Value (PPV), Negative Predictive Value (NPV) and Accuracy (ACC) were determined for each method. All methods were classified as significant (P < 0.05) predictors of pregnancy. Both ISG expression and PAG concentrations were greater (P < 0.05) in pregnant females than in non-pregnant and CL-NP females but did not differ (P > 0.05) from the PL group. ISG15 expression was greater (P < 0.05) in heifers than in cows, but this difference was not found in OAS1 expression and PAG concentrations. All the methods evaluated were proven to be adequate predictors of pregnancy, but greater accuracies were obtained through PAG concentrations and Doppler-US, due to the decreased number of false positive and false negative results.

Androstenedione and testosterone but not progesterone are potential biomarkers of pregnancy in Humpback Whales (Megaptera novaeangliae) approaching parturition.

The blubber steroid hormone profiles of 52 female humpback whales migrating along the east coast of Australia were investigated for seasonal endocrine changes associated with reproduction. Individuals were randomly sampled during two stages of the annual migration: before reaching the breeding grounds (northward migration; June/July), and after departing from the breeding grounds (southward migration; September/October). Assignment of reproductive status of the sampled individuals was based on season, single-hormone ranks and multi-variate analysis of the hormonal profiles. High concentrations of progesterone (>19 ng/g, wet weight), recognised as an indicator of pregnancy in this species, were only detected in one sample. However, the androgens, testosterone and androstenedione were measured in unusually high concentrations (1.6-12 and 7.8-40 ng/g wet weight, respectively) in 36% of the females approaching the breeding grounds. The absence of a strong accompanying progesterone signal in these animals raises the possibility of progesterone withdrawal prior to parturition. As seen with other cetacean species, testosterone and androstenedione could be markers of near-term pregnancy in humpback whales. Confirmation of these androgens as alternate biomarkers of near-term pregnancy would carry implications for improved monitoring of the annual fecundity of humpback whales via non-lethal and minimally invasive methods.

Use of a prediction method for early pregnancy status utilizing receiver operating characteristic curve analysis of peripheral blood leukocyte interferon-stimulated genes in Japanese-Black cattle.

A prediction method for early pregnancy status (pregnant or non-pregnant) in cattle that can be used within 3 weeks after insemination is desired. Interferon-stimulated genes (ISGs) in peripheral blood leukocytes (PBLs) have been examined as prediction molecules for determination of pregnancy status. Relative abundances of ISG15 and MX2 gene transcripts in PBLs were suitable biomarkers for the prediction of pregnancy status when there were assessments of Holstein cattle. In the present study, it was determined whether ISG biomarkers are applicable for predicting gestation in Japanese-Black (JB) cattle and evaluation of the applicability of receiver operating characteristic (ROC) analysis procedures for this purpose. There was assessment of the reliability of using average ISG values in PBLs collected during the estrous cycle (AVE) as a cutoff compared to the Youden index cutoff values. Application of AVE to assessment of pregnancy status in JB cattle indicated there was reliable predictions for pregnancy status when using ISG15 and MX2 values on day 21 after insemination, which coincided with the time of assessment in the previous study with Holstein cattle. The area under the curve values of the ROC curves confirmed the reliability of using ISGs to predict pregnancy from days 18 to 21 after insemination. Comparing AVE with Youden index values, there was confirmation of the accuracy of AVE for predicting gestation. The average mRNA transcript abundance values of ISG15 and MX2 may serve as excellent pregnancy biomarkers for cattle within 3 weeks of insemination.

False-Negative Urine Human Chorionic Gonadotropin Testing in the Clinical Laboratory.

BACKGROUND: Human chorionic gonadotropin (hCG) assays are used to detect pregnancy, and urine point-of-care tests are frequently used to triage patients. Under certain conditions, urine tests can fail to detect pregnancy, which can have serious consequences for patient management. OBJECTIVES: To understand the prevalence of different factors contributing to false-negative urinary hCG testing results at our institution. METHODS: Clinical data for patients with negative urine hCG results and subsequent positive or equivocal serum hCG results within a 1-year period were reviewed. RESULTS: Out of 9447 negative urine hCG results, 11 potential missed diagnoses were identified, with early gestational age as the most common factor, followed by ß-core hook effects. CONCLUSIONS: Although false-negative urine hCG test results are rare, understanding the commonly encountered reasons for inaccurate testing results can help clinical centers develop strategies to minimize risk for patients.