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Testicular steroids can alter the activity and expression of enzymes within the liver and may influence the metabolism of skatole and androstenone, which are responsible for boar taint. Plasma levels of estrone sulfate (E1S) are indicative of the steroidogenic capacity of the boar and are variable between animals of similar live weights at slaughter. This study aimed to characterize the relationship between steroidogenic capacity and the metabolism of boar taint compounds by relating plasma E1S levels at slaughter weight to the expression levels of genes regulating the metabolism of androstenone and skatole, along with their respective metabolite profiles. RT-qPCR was used to evaluate gene expression in the liver. Hepatocytes were also isolated and treated with androstenone or skatole, with metabolite levels in the incubation media quantified by high-performance liquid chromatography. Plasma E1S levels ranged from 2.2-108.5 ng/mL and were positively correlated with overall skatole metabolism (p = 0.038), the production of metabolites 3-methyloxindole (p = 0.026) and 3-hydroxy-3-methyloxindole (p = 0.036), and expression levels of key genes involved in skatole metabolism, specifically CYP2C33 (p = 0.0042), CYP2C49 (p = 0.022), and CYB5R1 (p = 0.017). There was no association between androstenone metabolism and plasma E1S concentrations; however, there was evidence of possible co-regulation amongst genes involved in the metabolism of androstenone, skatole, and estrogens. These findings indicate that steroidogenic capacity is related to the rate of skatole, but not androstenone metabolism, in slaughter-weight boars.

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Since 2022, European specifications for organic pig farming have evolved to distribute 100 % organic feed and to reinforce the link to the soil with feed resources that should primarily be obtained from the farm or the same region. Feeding strategy acts as a lever to improve various quality dimensions of organic (as well as conventional) pork, including intrinsic dimensions (carcass composition, nutritional, organoleptic, technological, sanitary qualities) and extrinsic dimensions related to animal farming (image). Diet may also influence the risk of undesirable odours or flavours that may be found in pork from non-castrated male pigs. This study aimed at evaluating the effects of a specific feeding strategy on several quality dimensions of organic meat from non-castrated male pigs. The experiment was conducted with 77 organic non-castrated male pigs (Piétrain NN × Large White) reared according to organic specifications and distributed in two batches. Within litters, male littermates were allocated at around 33 kg of live weight to either a Control group which received a Control feed (C) corresponding to the organic specifications or in a Bio+ group which received an organic test feed based mainly on French raw materials and which contained more fibres (faba bean and access to forages) and omega-3 fatty acids (linseed, camelina). All pigs were reared in the same building on deep straw bedding (1.3 m2/pig) with free outdoor access (1.0 m2/pig) using one pen per experimental group. Pigs were fed ad libitum until slaughter at about 125 kg live weight. Average daily gain, carcass weight and lean meat content did not differ significantly between C and Bio+ pigs. Compared to C, Bio+ pigs had higher (P < 0.05) ultimate pH in the loin (Longissimus muscle) and ham (Gluteus medius, and Semimembranosus) muscles, associated with a lower Longissimus muscle glycolytic potential (P < 0.001). Loin and ham meat from Bio+ vs. C pigs was lighter (P < 0.05) and had a more intense red colour (P < 0.10). The Bio+ strategy led to a lower Longissimus muscle n-6:n-3 fatty acid ratio (P < 0.001), indicating an improvement in pork nutritional value. Backfat skatole concentration was lower in Bio+ than in C pigs whereas backfat androstenone was higher in Bio+ than in C pigs (P < 0.05). Altogether, we demonstrated that the Bio+ strategy had positive impacts on several qualities of organic pork from non-castrated male pigs.

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Boar taint is an unpleasant odour found in the carcasses of entire male pigs, resulting from androstenone and skatole accumulation during pubertal development, and impacting pork quality. This study proposes the validation of an adapted chromatographic method for quantifying skatole and androstenone in the pigs' liquid fat using fluorescence detection. A good chromatographic separation was achieved, with skatole (SKA) and androstenone (AND) elution at 4.4 and 9.9 min., respectively. An external calibration method was applied, with calibration curves correlation coefficient of 0.9999 for both analytes. Detection limit values were 1.53 and 16.02 ng/g for SKA and AND, respectively. SKA recovery was 99.72±2.34 % (2.34 % RSD) and 102.84±1.62 % (1.57 % RSD) for AND. Results showed good precision values (repeatability <2.46 % RSD for SKA, <6.85 % RSD for AND; intermediate precision <2.87 % RSD for SKA, <6.98 % RSD for AND). The method's robustness was tested and the values were within the reference ranges. The validation results proved that the adaptation of an existing method resulted in good assessments of robustness, reliability and accuracy.

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Pork quality is determined by several attributes, among which odor and taste are the utmost significant. Therefore, this study was aimed to assess the effects of boar odor hormone concentration on the quality traits and sensory acceptability of pork. A total twenty-six (26) non-castrated 3-way crossbred (Landrace × Yorkshire × Duroc) pigs were selected with an average body weight (ABW) 115.6 kg before to slaughter. The three treatment groups (low, medium and high) were divided according to the androstenone concentration. In experiment 1, for meat quality traits carcass was selected based on androstenone concentration: low (LC, 0.64-0.69 µg/g, n = 9), medium (MC, 0.70-0.99 µg/g, n = 7) and high (HC, 1.00-1.69 µg/g, n = 10). In experiment 2, for sensory evaluation carcasses were also selected based on the abovementioned conditions. Results revealed that androstenone concentration not effect on proximate components, meat quality traits and fatty acids except palmitoleic acid. Sensory evaluation data showed that boar taint and meat boar taint were significantly increased in a concentration-dependent manner from low to high, whereas, gravy and meat flavor preference were significantly increased in LC group than HC group. In addition, correlation analysis showed that boar taint and meat boar taint were positively, and gravy and meat flavor preference were negatively correlated with boar taint hormones. In essence, our findings indicate that androstenone concentration had no effect on meat qualities, but a high concentration of androstenone had a negative effect on the sensory characteristics in uncastrated pigs.

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This study aims to characterize a complete volatile organic compound profile of pork neck fat for boar taint prediction. The objectives are to identify specific compounds related to boar taint and to develop a classification model. In addition to the well-known androstenone, skatole and indole, 10 other features were found to be discriminant according to untargeted volatolomic analyses were conducted on 129 samples using HS-SPME-GC×GC-TOFMS. To select the odor-positive samples among the 129 analyzed, the selection was made by combining human nose evaluations with the skatole and androstenone concentrations determined using UHPLC-MS/MS. A comparison of the data of the two populations was performed and a statistical model analysis was built on 70 samples out of the total of 129 samples fully positive or fully negative through these two orthogonal methods for tainted prediction. Then, the model was applied to the 59 remaining samples. Finally, 7 samples were classified as tainted.

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In this work, we studied the backfat transcriptome of surgically castrated (C), intact (I) and intact fed an experimental diet (IE) outdoor-reared male Alentejano (AL) pigs. The experimental diet was a high-fiber diet with locally produced legumes and by-products associated with a boar taint reduction effect. At slaughter (~160 kg), backfat samples were collected for total RNA sequencing. Intact pigs presented leaner carcasses, more total collagen, and more unsaturated intramuscular fat content than C animals. A total of 2726 differentially expressed genes (DEGs, |log2 FC|> 0.58, q < 0.05) were identified between C and I with overexpressed genes related to muscular activity (MYH1, ACTA1) or collagen metabolism (COL1A1, COL1A2) in I pigs. Between C and IE, 1639 DEGs of genes involved in lipidic metabolism (LEP, ME1, FABP4, ELOVL6) were overexpressed in C. Finally, only 28 DEGs were determined between I and IE. Clustering results indicated a drastic influence of the testis in the transcriptome of subcutaneous fat of AL pigs, while the diet had a marginal effect. Diet can reduce stress by increasing satiety in animals, and could have induced an increase of skatole degradation due to the higher expression of the CYP2A19 gene in the IE group.

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Immunocastration and rearing of entire males (EMs) are sustainable alternatives to surgical castration. However, these animal carcasses have variable risk of boar taint and should be identified at the slaughter line. We aimed to identify a simple and reliable indicator of androstenone-related boar taint by evaluating pelvic urogenital tract weight as a marker of boar-taint animals at the slaughter line. The pelvic urogenital tract, testes, and accessory sex glands of EMs and immunocastrates (ICs) were collected, dissected, and weighed, before colorimetric measurements of testicular tissue. Additionally, GnRH antibody titers and testosterone, androstenone, and skatole levels were determined. Our results showed that 81.8% of EMs had androstenone levels above the risk threshold (>0.5 µg/g fat; EM/Ahigh subgroup), whereas in ICs, the C/Ahigh subgroup with androstenone >0.5 µg/g fat accounted for only 4.3%. Androstenone levels correlated negatively with GnRH antibody titers and positively with testosterone levels and reproductive organ weights. Identification of ICs with androstenone levels above the threshold (IC/Ahigh subgroup) may be achieved via testes or pelvic urogenital tract weight measurements. However, in EMs, the latter is a more reliable parameter. A principal component analysis based on these variables and hierarchical clustering also distinguished the Ahigh from the Alow subgroup, irrespective of IC/EM. The findings highlight the possible use of pelvic urogenital tract weight along with testes weight as a simple, reliable, and efficient morphometric indicator for identifying androstenone-positive carcasses of different sex categories.

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Androstenone, a volatile steroid that possesses pheromonal activity, is responsible for boar taint, sexual interactions, and reproduction in pigs. A wide range of analytical methods has been developed to quantify and detect androstenone in adipose tissue and blood, which are invasive procedures. Therefore, the present study aimed to develop a non-invasive method to detect and quantify the androstenone. We produced group-specific polyclonal androstenone antibody to standardize and validate an enzyme immunoassay to measure faecal and urinary androstenone in Yorkshire boars and sows. Parallelism was performed to determine the immunoreactivity between faecal and urinary immunoreactive androstenone and respective antibody. In boars, urinary and faecal androstenone concentrations were higher on the day of mounting and copulation with sows. In sows, we also measured faecal progesterone metabolites to confirm the oestrus and mating. Faecal androstenone concentrations were peaked on the day of oestrus and mating in sows. Our results suggest that androstenone could be detected and quantified in faecal and urine samples of boars and sows. •Developed an enzyme immunoassay for measuring 5α-androst-16-en-3-one as a marker of boar taint and sex pheromone in urine and faeces of pigs•Detection of 5α-androst-16-en-3-one using a non-invasive method.

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Reduced-fat cured sausages were evaluated as a strategy to reduce boar taint in entire male pork products with high levels of androstenone and skatole, both lipophilic. Three fuet-type sausages (two replicates each) were developed: the control (C) (60% lean, 33.69% fat), and two reduced-fat (60% lean 21.19% fat) products; R1, 6% inulin, 0.5% ß-glucan and R2, 3% inulin, 0.5% grape skin and 1% ß-glucan. All of them were prepared from entire male pork with an androstenone concentration of 6.887 µg/g and 0.520 µg/g of skatole. Significant differences (p ≤ 0.001) in the moisture content were described between the fuet R1 and the C and R2, which obtained the highest percentage. Regarding the CIELAB, the C samples had the highest L* value, while the R2 sausages were the darkest. Boar taint was reduced in both R1 and R2, with a greater reduction in R2 (p ≤ 0.000). The addition of inulin and ß-glucan in fuet R1 provided a similar technological and sensory profile to C. However, both strategies provided a reduction of sexual odour, which was higher when grape skins were included. In addition, R2 gave a characteristic sausage with more odour and flavour, dark colour and overall rating than C and R1.

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The 2-oxidation, 3-methyl hydroxylation, and 6-hydroxylation of skatole (a contributor to boar taint) mediated by minipig liver microsomes and recombinant P450 enzymes expressed in bacterial membranes were investigated.At low substrate concentrations of 10 µM, the formation rates of indole-3-carbinol, 6-hydroxyskatole, and the sum of 3-methyloxindole, indole-3-carbinol, and 6-hydroxyskatole in male minipig liver microsomes were significantly lower than those in female minipig liver microsomes.Compensatory 3-methyloxindole and indole-3-carbinol formation in minipig liver microsomes, which lack 6-hydroxyskatole formation in males, was mediated partly by liver microsomal P450 1A2 and P450 1A2/2E1, respectively. These enzymes were suppressed by typical P450 inhibitors in female minipig liver microsomes.Among the 14 pig P450 forms evaluated, P450 2A19 was the dominant form mediating 3-methyloxindole, indole-3-carbinol, and 6-hydroxyskatole formation from skatole at substrate concentrations of 100 µM. Positive cooperativity was observed in 3-methyloxindole formation from skatole mediated by male minipig liver microsomes and by pig P450 3A22 with Hill coefficients of 1.2-1.5.These results suggest high skatole 2-oxidation, 3-methyl hydroxylation, and 6-hydroxylation activities of pig P450 2A19 and compensatory skatole oxidations mediated by pig P450 1A2, 2E1, or 3A22 in male minipig liver microsomes.

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Boar taint masking strategies were developed using hydrocolloids and spices to produce edible gels and films. Carrageenan (G1) and agar-agar (G2) were used for the gels and gelatin (F1) and alginate+maltodextrin (F2) for the films. The strategies were applied to both castrated (control) and entire male pork with high levels of androstenone and skatole. The samples were evaluated sensorially by a trained tasting panel through quantitative descriptive analysis (QDA). Lower hardness and chewiness of entire male pork, with high levels of boar taint compounds object of study, were found with the carrageenan gel, due to the better carrageenan gel adherence to the loin. The films showed that the gelatin strategy tended to have a certain "sweet" taste, and a higher overall masking than the alginate+maltodextrin film. In conclusion, a trained tasting panel found that gelatin film masked boar taint the most, followed by the alginate+maltodextrin film and the carrageenan-based gel.

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This study aimed at understanding which molecules were responsible for the differences existing in boar taint sensory evaluation. The latter was therefore linked to the results of skatole and androstenone chemical analyses, fatty acid composition and VOC profiles of heated backfat. This study confirmed that some discrepancy exists between chemical analysis and sensory evaluation of tainted backfats. Significant correlations between human nose scores and fatty acid composition were not revealed. Strong correlations between emissions and contents in skatole and androstenone were found. Oxidation products of polyunsaturated fatty acids, with fatty odor descriptors, were found to be more present in the VOC profiles of boar fat considered untainted through the human nose methodology. Weak coefficient of determination for partial least square regression indicates that other factors, yet unknown, are responsible for sensory evaluation outcomes. These findings hence support the idea that high human nose score is mainly due to boar taint compounds rather than general differences in VOC profiles.

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Traditional outdoor pig farming is renowned for its emphasis on animal welfare and the production of highly valued, quality meat. While seasonality is known to impact certain animals, particularly those raised outdoors, there is a lack of research on Bísaro boars, a native Portuguese breed. This research study was conducted on a total of 20 male entire Bísaro pigs, reared in outdoor pens from 4 to 13 months old, and subsequently slaughtered. The animals were divided into two groups: one slaughtered in winter (Wi, n = 9), and the other in summer (Su, n = 11). The objective was to evaluate testicular morphometry, boar taint compounds, and meat quality traits, including sensory analysis and fatty acid profile. Testicles from the Su group exhibited reduced volume, indicating diminished functionality during that season. While no significant differences were observed in the boar taint compound analysis, panelists could discern a more intense aroma and flavor of boar taint in the Su meat. Other meat quality traits showed no significant variations, but the fatty acid profile displayed higher values in the Wi group. This study reveals that Bísaro boars experience reproductive seasonality, leading to variations in boar taint compounds across the seasons. This information is crucial for farm planning.

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Mass spectrometry (MS)-based techniques, including liquid chromatography coupling, shotgun lipidomics, MS imaging, and ion mobility, are widely used to analyze lipids. However, with enhanced separation capacity and an optimized chemical derivatization approach, comprehensive two-dimensional gas chromatography (GC×GC) can be a powerful tool to investigate some groups of small lipids in the framework of lipidomics. This study describes the optimization of a dedicated two-stage derivatization and extraction process to analyze different saturated and unsaturated fatty acids in plasma by two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) using a full factorial design. The optimized condition has a composite desirability of 0.9159. This optimized sample preparation and chromatographic condition were implemented to differentiate between positive (BT) and negative (UT) boar-tainted pigs based on fatty acid profiling in pig serum using GC×GC-TOFMS. A chemometric screening, including unsupervised (PCA, HCA) and supervised analysis (PLS-DA), as well as univariate analysis (volcano plot), was performed. The results suggested that the concentration of PUFA ω-6 and cholesterol derivatives were significantly increased in BT pigs, whereas SFA and PUFA ω-3 concentrations were increased in UT pigs. The metabolic pathway and quantitative enrichment analysis suggest the significant involvement of linolenic acid metabolism.

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"Boar taint" compounds influence the sexual behavioral responses of sows and stimulate their reproduction. This paper reports a fast, easier, and a non-invasive analytical method for the analysis of three "boar taint" compounds in boar' saliva samples: androstenone, androsten-3α-ol, and androsten-3ß-ol. This method was developed and validated based on solid-phase extraction (SPE) and multidimensional gas chromatography-mass spectrometry (MDGC-MS). All the compounds were detected without derivatization. This method affords good reproducibility (4%-8%), accuracy (80%-105%), precision (5.5%-9.1%), linearity (R2 = 0.98-0.99), and lower limits of quantitation (LLOQ) (0.1-0.2 µg/L). Although the presence of these compounds in saliva has been known for a long time, no simple and easy analytical method has been developed.

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The nuclear receptors PXR, CAR, and FXR are activated by various ligands and function as transcription factors to control the expression of genes that regulate the synthesis and metabolism of androstenone and skatole. These compounds are produced in entire male pigs and accumulate in the fat to cause the development of a meat quality issue known as boar taint. The extent of this accumulation is influenced by the synthesis and hepatic clearance of androstenone and skatole. For this reason, PXR, CAR, and FXR-mediated signaling pathways have garnered interest as potential targets for specialized treatments designed to reduce the development of boar taint. Recent research has also identified several metabolites produced by gut microbes that act as ligands for these nuclear receptors (e.g., tryptophan metabolites, short-chain fatty acids, bile acids); however, the connection between the gut microbiome and boar taint development is not clear. In this review, we describe the nuclear receptor signaling pathways that regulate the synthesis and metabolism of boar taint compounds and outline the genes involved. We also discuss several microbial-derived metabolites and dietary additives that are known or suspected nuclear receptor ligands and suggest how these compounds could be used to develop novel treatments for boar taint.

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Painful castration of male piglets to avoid boar taint can potentially be replaced by three more ethical alternatives: entire male production in combination with a detection method, immunocastration (an active vaccination against the gonadotrophin-releasing factor, GnRF), and castration with pain relief (anesthesia and/or analgesia). With the aim of abandoning piglet castration and facilitating internal trade, the European Union (EU) was initially in favor of a single alternative. Immunocastration was proposed as a potential solution, but it has not yet been sufficiently assessed regarding its market potential. To address this point, this paper uses scenario analysis to examine whether and under what conditions immunocastration could be the general solution sought by the EU. The study constructs two extreme scenarios: one in which all uncertain elements negatively influence the growth of immunocastration; another in which all uncertain elements have positive influences. These scenarios provide insights into the variance in possible futures for the implementation of immunocastration. The results show that it is unlikely that immunocastration will become a single solution for all producers in the EU, because it is not the optimal solution for all types of EU pork production systems (i.e., cost-efficiency oriented, quality oriented, animal-friendly oriented, import dependent). Rather than debating and looking for evidence about which single method is the best for the entire EU, EU authorities are advised to allow the co-existence of all alternatives and to develop protocols for applying them in the pork industry.

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The practice of surgical castration of piglets and its alternatives is still under debate. Production of boars may impair meat quality due to boar taint and reduced tenderness compared to meat from surgically castrated male pigs, while immunocastration reduces boar taint and may improve meat quality but seems to be less accepted by the pig chain. In this study, we aimed to evaluate the consumer's sensory appreciation of meat from barrows (BAs), immunocastrates (ICs) and boars (BOs) in six European countries, taking into account the selection of tainted carcass and consumers' appreciation of boar taint. Loin chops of 30 BAs, 30 ICs and 30 BOs were evaluated by 752 consumers in six countries: Belgium, Czech Republic, Poland, Portugal, Romania and Spain. Consumers rated odour, flavour, tenderness, juiciness, overall liking and willingness to buy and sensitivity to and liking of androstenone (AND) and liking of skatole (SKA) was also tested. In each of the six countries, consumers liked the odour of the BO samples less than that of BA, and IC intermediate. For flavour, tenderness, juiciness, overall liking and willingness to buy, liking scores given by the Czech, Polish and Portuguese consumers significantly differed between the BA, BO and IC. Willingness to buy was highest for BA by Czech and Polish consumers and for BA and IC by Portuguese consumers. The frequency of the negative check all terms that apply terms also differed, with a higher frequency of disgusting for BO compared to BA and IC and of off-flavour, irritating, manure, sweat, disappointing compared to BA, and intermediate for IC. 31% of the consumers disliked the odour of AND (NEGAND), and 36% of them were not sensitive; in contrast, 77% of the consumers disliked SKA (NEGSKA). The decrease in flavour liking score for BO compared to BA and IC was more outspoken by the NEGAND consumer, while NEGSKA consumers gave an overall lower liking score independent of the type of male pig. The results of this study indicate that IC can be a valid alternative for surgical castration.

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With the increasing age of boars, the possibility of androstenone (5α-androst-16-en-3-one (AND), 5α-androst-16-en-3α-ol (α-AND), and 5α-androst-16-en-3ß-ol (ß-AND)) occurrence increases as well. The aim of this study was to evaluate concentrations of androstenone compounds in the saliva of boars concerning the age of animals. In total, 72 boars were evaluated (24 boars per replication). The effect of age (three different ages-152, 163, and 172 days) was observed, and (Landrace × Large White) × Pietrain genotypes were used. Chemical analysis of saliva samples was conducted by multidimensional gas chromatography/mass spectrometry (MDGC/MS). Salivary α-AND increased with age (p < 0.05), and positive correlations were found between age and submaxillary salivary gland weight (p < 0.05), age, and salivary AND concentration (p < 0.05), body weight and submaxillary salivary gland weight (p < 0.05), submaxillary salivary gland weight and salivary ß-AND concentration (p < 0.05), as well as submaxillary salivary gland weight and total salivary 5α-androstenone (AND total) concentration (p < 0.001). Nowadays, animal welfare is becoming a more and more discussed topic, and pig breeding is not an exception. Specifically, the castration ban is a current issue, so it is very important to know as much as possible about compounds responsible for boar taint. Androstenone's appearance in boars' saliva could be one of the most important precursors for future early detection of boar taint.

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Introduction: Most male pigs are surgically castrated to avoid puberty-derived boar taint and aggressiveness. However, this surgical intervention represents a welfare concern in swine production. Disrupting porcine KISS1 is hypothesized to delay or abolish puberty by inducing variable hypogonadotropism and thus preventing the need for castration. Methods: To test this hypothesis, we generated the first KISS1-edited large animal using CRISPR/Cas9-ribonucleoproteins and single-stranded donor oligonucleotides. The targeted region preceded the sequence encoding a conserved core motif of kisspeptin. Genome editors were intracytoplasmically injected into 684 swine zygotes and transferred to 19 hormonally synchronized surrogate sows. In nine litters, 49 American Yorkshire and 20 Duroc liveborn piglets were naturally farrowed. Results: Thirty-five of these pigs bore KISS1-disruptive alleles ranging in frequency from 5% to 97% and did not phenotypically differ from their wild-type counterparts. In contrast, four KISS1-edited pigs (two boars and two gilts) with disruptive allele frequencies of 96% and 100% demonstrated full hypogonadotropism, infantile reproductive tracts, and failed to reach sexual maturity. Change in body weight during development was unaffected by editing KISS1. Founder pigs partially carrying KISS1-disruptive alleles were bred resulting in a total of 53 KISS1 +/+, 60 KISS1 +/-, and 34 KISS1 -/- F1 liveborn piglets, confirming germline transmission. Discussion: Results demonstrate that a high proportion of KISS1 alleles in pigs must be disrupted before variation in gonadotropin secretion is observed, suggesting that even a small amount of kisspeptin ligand is sufficient to confer proper sexual development and puberty in pigs. Follow-on studies will evaluate fertility restoration in KISS1 KO breeding stock to fully realize the potential of KISS1 gene edits to eliminate the need for surgical castration.