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Article

Innovation on Swine Semen Storage: Bacteriostatic Coating vs. Conventional Blister in Commercial Swine Semen Production

by
Janine de Camargo
1,2,
Pedro Nacib Jorge-Neto
3,4,*,
Érika Lopes Madruga
5,
Maria Gessica Daniel de Oliveira
5,
Gilson Fruhling
5,
José Victor Braga
6,
Rosangela Poletto
2 and
Ricardo Zanella
1,*
1
Escola de Ciências Agrarias, Inovação e Negócios, Universidade de Passo Fundo, Passo Fundo 99042-800, RS, Brazil
2
Instituto Federal de Educação, Ciência e Tecnologia do Rio Grande do Sul (IFRS), Campus Sertão, Sertao 99170-000, RS, Brazil
3
Faculty of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo 05508-220, SP, Brazil
4
Faculty of Veterinary Medicine and Husbandry, Federal University of Mato Grosso do Sul, Campo Grande 79070-900, MS, Brazil
5
Nutribras Alimentos, Vera 78890-000, MT, Brazil
6
Laboratório de Reprodução Animal, Universidade Federal de Pelotas, Pelotas 96010-610, RS, Brazil
*
Authors to whom correspondence should be addressed.
AgriEngineering 2025, 7(10), 338; https://doi.org/10.3390/agriengineering7100338
Submission received: 16 July 2025 / Revised: 28 September 2025 / Accepted: 29 September 2025 / Published: 10 October 2025
(This article belongs to the Special Issue Advancing Livestock Production: Management Strategies and New Technologies)
Review Reports Versions Notes

Abstract

This study investigated the effectiveness of a bacteriostatic-coated blister in preserving swine semen quality and its impact on reproductive performance. Two experiments were conducted: an in vitro assessment of the blister’s bacteriostatic efficacy and semen quality during three days of storage (Experiment 1), and a seven-day commercial farm trial evaluating its effect on reproductive outcomes in artificially inseminated gilts and sows (Experiment 2). In Experiment 1, the bacteriostatic blister effectively controlled bacterial proliferation, maintaining counts below 2 log10, comparable to controls with added antibiotics. Sperm quality parameters, including total and progressive motility, consistently exceeded the critical threshold for artificial insemination. Experiment 2 demonstrated that the bacteriostatic coating did not negatively affect key reproductive performance indicators, such as farrowing rate, total piglets born, or live piglets under commercial conditions. These findings suggest that the bacteriostatic-coated blister offers a viable, potentially antibiotic-free, alternative for semen preservation, extending storage viability for up to seven days. This technology supports sustainable reproductive practices, representing a significant advancement in commercial swine production.

1. Introduction

In commercial swine production, artificial insemination (AI) is employed in over 90% of females and is pivotal to economic success [1]. This technique is continuously refined to improve reproductive outcomes [2,3]. AI success hinges on multiple factors related to both the sow and the boar, as gamete quality directly affects embryonic and fetal development [4,5]. Consequently, evaluating the fertilization capacity of boar semen is crucial for optimizing AI in swine production systems [6].
Semen doses are typically stored at temperatures between 15 °C and 18 °C, reflecting the physiological sensitivity of swine spermatozoa. Lower temperatures are avoided due to their detrimental effects on sperm viability [7,8,9]. However, this temperature range can promote bacterial growth [10], which degrades semen quality and facilitates pathogen transmission to inseminated females, negatively impacting fertility and litter size [3,11].
The addition of antibiotics to semen extenders is a standard industry practice to mitigate bacterial contamination. Nevertheless, the indiscriminate use of antibiotics has led to the global emergence of resistant microorganisms [12,13,14], posing a significant challenge to global public health [15]. Given the global relevance of pork production, over 10 million liters of semen with antibiotic extenders are utilized annually in swine AI [16].
The pursuit of sustainable practices in pig farming underscores the need to balance animal production, economic viability, and environmental sustainability, as emphasized by the One Health concept [17,18]. In this context, research has explored alternative strategies to reduce bacterial contamination in semen without relying on antibiotics. These methods range from improving hygiene during semen collection, processing, and storage [16] to using antimicrobial supplements, lowering storage temperatures [8,19,20], or applying colloid centrifugation [21,22,23,24]. Each approach, however, faces distinct challenges. Storing semen below 10 °C can compromise sperm quality by altering membrane structure and permeability [25,26]. Although colloid centrifugation is an effective antibiotic-free method for physically removing bacteria, its broader implementation in a commercial boar stud is hindered by economic and logistical constraints, underscoring the critical need to align biological efficacy with practical and financial viability [21,22].
Preserving sperm quality and ensuring reproductive success remain critical challenges in the swine industry, highlighting the necessity to develop technologies that reduce reliance on antimicrobials in semen extenders [3]. The aim of this study was, therefore, to evaluate the capacity of a bacteriostatic-coated blister to preserve sperm quality during storage and its impact on reproductive performance, both in vitro and in a commercial swine production setting, with a primary focus on confirming that the incorporated bacteriostatic agent did not adversely affect these parameters.

2. Materials and Methods

2.1. Ethical Note

This study received approval from the Animal Use Ethics Committee of the University of Passo Fundo, Brazil (CEUA-UPF), under protocol No. 005/2023.

2.2. Study Strategy

Our study was conducted in two independent trials. The first experiment, performed at the Reprogene Lab of the University of Passo Fundo, focused on the in vitro validation of the bacteriostatic efficacy of the BactiBag® blister (Ref. 025881; IMV Technologies, L’Aigle, France) and the evaluation of semen quality stored within it.
The second trial assessed the impact of the BactiBag® blister on the reproductive performance of gilts and sows subjected to artificial insemination at a commercial swine production site (Swine Farm Nutribrás Alimentos, Vera, MT, Brazil; 12°19′17.7′′ S 55°26′41.9′′ W). This trial also evaluated the blister’s efficacy in maintaining semen quality during storage.

2.3. Procedures

2.3.1. Semen Quality Assessment-Protocol in Computer-Assisted Semen Analysis

Semen quality parameters were assessed using the Computer-assisted Semen Analysis (CASA-IVOS II, Hamilton Thorne, Beverly, MA, USA) system. For the analysis, freshly diluted samples were first pre-incubated for 10 min at 37 °C to allow for thermal equilibration. The CASA assessment was then performed at the same temperature of 37 °C. The parameters analyzed included total sperm motility [defined as curvilinear velocity (VCL) > 24 μm/s and lateral head displacement (ALH) > 1 μm], progressive motility, and various movement characteristics. These characteristics encompassed lateral head displacement amplitude (ALH), beat cross frequency (BCF, Hz), curvilinear velocity (VCL, μm/s), average path velocity (VAP, μm/s), straight-line velocity (VSL, μm/s), straightness (STR, VSL/VAP, %), and linearity (LIN, %).
For analysis, each semen sample was gently homogenized, and a 3 µL aliquot was placed in the chamber of a Leja slide (Ref. 025107, IMV Technologies) tilted at 45°, with the exterior blotted dry. To ensure sample stability, six to eight successive fields along the slide’s central axis were analyzed using the CASA system (IVOS II, Hamilton Thorne, Beverly, MA, USA). The Animal Breeders II software (version 1.13.7; Hamilton Thorne, Beverly, MA, USA), configured for pigs, was used according to the manufacturer’s recommendations. The “Auto-First Frame” option ensured proper marking of sperm tails and heads. Analyses were recorded at 60 frames per second using 100× magnification, with a minimum of 400 spermatozoa analyzed per sample.

2.3.2. Experiment 1-Bacteriostatic Efficacy of the Bacteriostatic Coating Blister and In Vitro Quality Assessment of Semen Dose Storage

This experiment utilized two sets of animals, totaling 11 males commercial hybrids (Landrace x Large White), all housed in the same facilities. The first set comprised six boars, while the second included five animals, each with an average age of 13.3 ± 3.8 months. All boars were individually housed in temperature-controlled pens (9 m2 per boar) maintained between 16 and 18 °C. They had ad libitum access to water and received a daily diet of 2.5 kg of a corn and soybean-based commercial feed. All boars had known fertility profiles and were routinely subjected to semen collection. In Experiment 1, the analysis of bacteriostatic efficacy was performed with a total of 11 ejaculates, while the detailed evaluation of semen quality (including motility and kinematics) was conducted on a subset of 5 of these ejaculates. This approach was adopted to ensure a robust sampling of the bacteriostatic coating’s ability to control bacterial proliferation across a larger number of samples, while accounting for the natural variability of ejaculates. The in-depth assessment of sperm parameters was concentrated on a subgroup for a more detailed evaluation of sperm functionality, ensuring the viability and consistency of the results. Normospermic ejaculates were collected from 11 males in two replicates, spaced seven days apart, using the double-gloved hand technique to provide a double layer of protection from contamination. A total of 11 ejaculates were collected for the study. Immediately after sampling, sperm quality parameters, including motility and concentration, were assessed using the CASA system (IVOS II, Hamilton Thorne, Beverly, MA, USA). Only normospermic ejaculates with at least 70% motility and a maximum of 20% abnormal sperm cells were selected for the experiment.
Quality-approved ejaculates from all boars were diluted to a concentration of 2.5 × 109 spermatozoa in a total volume of 90 mL.
Ejaculates from each boar were manually extended in Beltsville Thawing Solution (BTS), with or without the addition of 0.25 mg/mL gentamicin sulfate (Sigma Aldrich, St. Louis, MO, USA; Ref. G1264). The BTS semen extender formula included 205 mM glucose (C6H12O6), 20.4 mM sodium citrate (Na3C6H5O7), 10.0 mM potassium chloride (KCl), 15 mM sodium bicarbonate (NaHCO3), and 3.36 mM ethylenediaminetetraacetic acid disodium salt dihydrate (EDTA), with the pH adjusted to 7.2 [27].
Diluted ejaculates from each male were subjected to four different treatment conditions:
(a)
Bacteriostatic Coating Blister Plus Antibiotic: Doses were diluted in BTS extender at 34 °C, supplemented with 0.25 g/L gentamicin sulfate, and stored in a blister with a bacteriostatic coating.
(b)
Bacteriostatic Coating Blister Without Antibiotic: Doses were diluted in BTS extender at 34 °C without antibiotics and stored in a blister with a bacteriostatic coating.
(c)
Control with Antibiotic: Doses were diluted in BTS extender at 34 °C, supplemented with 0.25 g/L gentamicin sulfate, and packaged in conventional GTB BAG II blisters (Ref 022331; IMV Technologies, L’Aigle, France).
(d)
Control Without Antibiotic: Doses were diluted in BTS extender at 34 °C without antibiotics and packaged in conventional blisters.
These diluted ejaculates were then stored between 16 °C and 18 °C.
Bacteriostatic Efficacy of BactiBag®
Bacterial proliferation was evaluated in samples from 11 boar ejaculates subjected to four different treatment conditions (n = 44 total samples), as described in Section 2.2. The assessment was conducted after 72 and 120 h of storage at 17 °C.
To quantify the bacterial load, serial dilutions (1:10, 1:100, and 1:1000) of each semen sample were prepared in sterile Phosphate-Buffered Saline (PBS; 0.1 M, pH 7.3). For plating, a 1.0 mL aliquot from each dilution was spread in triplicate onto Tryptic Soy Agar (TSA) plates. Following incubation at 37 °C for 48 h, colonies were counted. The final bacterial concentration was calculated from the average of the three plates from a countable dilution, expressed as colony-forming units per milliliter (CFU/mL), and then log-transformed for analysis (Log CFU/mL).
In Vitro Quality Assessment of Semen Dose Storage
Semen samples (n = 40) for this evaluation were derived from five ejaculates, one from each of the five boars in the second animal set (as previously described). These samples, processed as detailed in Section 2.2, were stored at a controlled temperature of 16 to 18 °C. Total motility, progressive motility, and sperm Kinematics were assessed using the CASA system (detailed in Section 2.1) at 72 and 120 h post-collection.

2.3.3. Experiment 2–Field Assessment of Bacteriostatic Coating Blister

This experiment aimed to evaluate the reproductive performance of gilts and sows subjected to AI using semen stored in blisters with a bacteriostatic coating on a commercial farm. To replicate commercial conditions, the insemination doses were produced using a commercial extender containing antimicrobials. Additionally, the study assessed the efficacy of these blisters in maintaining semen quality for up to 168 h under farm conditions.
Preparation of Insemination Doses
Sixty mature boars (Landrace, n = 7; Large White, n = 10; Duroc, n = 43), aged between one and three years and housed at the Nutribrás Genetic Diffusion Unit in Mato Grosso, Brazil, were utilized. Ejaculates were collected weekly over four consecutive weeks using the gloved-hand technique, with four-day intervals between collections, consistent with standard farm practice. Ejaculates were directed into 3.6-L semen collection bags equipped with filters (Ref DZA000195, IMV Technologies, Campinas, SP, Brazil) and analyzed for sperm quality using the CASA system (IVOS II), following the protocol described in Section 2.1.
Individual ejaculates (1 to 4 per boar) were isothermally diluted using NutriXcell Ultra extender (IMV Technologies, L’Aigle, France) maintained at 37 °C. This process yielded a total of 153 processed ejaculates, distributed as follows: 15 from Landrace, 19 from Large White, and 119 from Duroc.
Following dilution, each ejaculate was divided into two experimental blister treatments: (1) blisters with a bacteriostatic coating and (2) conventional blisters. Blisters were filled using two GTB 1000V2 machines (IMV Technologies, L’Aigle, France). For post-cervical artificial insemination (PCAI), semen doses were adjusted to 40 mL, containing 1.25 × 109 spermatozoa per dose. For conventional insemination, doses were 80 mL, containing 2.2 × 109 spermatozoa per dose. All doses were stored at controlled temperatures between 16 °C and 18 °C and used for AI within 24 to 48 h post-preparation, as directed by farm management. Additionally, one sample bag from each treatment and volume type (totaling 4 samples per ejaculate) was retained for further storage analyses, resulting in up to 16 samples per boar over the collection period.
Storage Efficacy of Bacteriostatic Coating Blister on Semen Quality
Semen samples from both bacteriostatic coating blisters and conventional blisters were analyzed for total motility, progressive motility, and sperm Kinematics. These assessments were performed at 24, 72, 120, and 168 h post-collection using the CASA system, as previously described in Section 2.1. Samples collected at time zero (0, control), considered fresh semen, were analyzed for all boars as outlined in Section 2.1.
Sperm agglutination was additionally assessed following the methodologies described by Maroto Martín et al. [11] and Gączarzewicz et al. [28]. For each storage time point (excluding time zero due to analysis limitations on commercial farms), three microliters of semen were examined under a phase contrast microscope across at least eight fields. The degree of agglutination was categorized on a scale from 0 to 5:
  • 0: No agglutination
  • 1: 5–10% agglutination
  • 2: 10–20% agglutination
  • 3: 20–30% agglutination
  • 4: 30–40% agglutination
  • 5: 40–50% agglutination
Clusters of nonmotile spermatozoa were not considered during the agglutination analysis.
Reproductive Performance-Insemination, Pregnancy Diagnosis, and Birth
A field insemination trial was conducted at a pig farm in Vera-MT, Brazil, to compare the reproductive performance of semen stored in blisters with bacteriostatic coating to that in conventional blisters. All semen was stored at 17 °C.
The 2192 commercial hybrid females (696 gilts and 1496 sows) participating in the study were housed in temperature-controlled gestation facilities (16–18 °C). Each animal had ad libitum access to water and received 2.5 kg of a corn and soybean-based commercial diet daily. During the AI, sows and gilts were kept individually housed on pens with 60 cm × 2.20 m of area per female. The pregnant animals were subsequently moved to a group house, providing at least 3.0 m2 per female.
Females were grouped by parturition category and underwent the artificial insemination (AI) protocol. Each week for four consecutive weeks, 174 gilts and 374 multiparous sows (two to eight parities) were inseminated. Half of these received semen doses stored in bacteriostatic-coated blisters, while the other half received doses in conventional blisters. This resulted in a total of 696 gilts (parity 0) and 1496 sows (parity 2 to 8) included in the study. Estrus detection of females in both categories was performed twice daily using the back-pressure test (BPT) in the presence of a boar. Each female received on average three consecutive inseminations at 12-h intervals, consistently using the same type of blister. Semen doses were stored at 16–18 °C for 48 h before insemination. The females (sows and gilts) were kept in individual stalls, during the AI. They remained in these stalls until pregnancy was confirmed. Only after the pregnancy diagnosis were confirmed females were transferred to group housing. Pregnancy checks were performed 30 days post-insemination via ultrasonography (SC Tecnoscan, IMV Imaging, Bellshill, UK).
Approximately one week before farrowing, pregnant females were transferred to the farrowing area, where they were housed in partitions with 24-h monitoring in farrowing crates. Reproductive performance data, including pregnancy rate, farrowing rate, total piglets born, live-born piglets, and total litter weight, were collected for each female to allow for detailed analysis under varying AI experimental conditions.

2.4. Statistical Analysis

For Experiment 1, bacterial contamination and sperm parameters were compared across different groups (Blisters and Antibiotic) and collection time points (72 and 120 h). Repeated measures ANOVA was utilized to account for within-subject variability over time, allowing for the assessment of main effects and interactions between group membership and time points. When significant effects were identified, Tukey’s post-hoc test was employed to determine specific differences.
In Experiment 2, statistical analyses assessed the effects of storage bag type (bacteriostatic coating blister vs. conventional blister) on sperm motility, sperm Kinematics, agglutination, and reproductive performance. The influence of bag type, storage duration, storage volume, and boar breed, along with the interactions were tested.
Repeated measures ANOVA was performed to assess the effects of bag type, storage duration, and storage volume on total and progressive sperm motility. Tukey’s post-hoc test was applied for specific differences when significant main effects were detected. Interaction effects between bag type and other variables (storage duration and volume) were also examined.
Furthermore, sperm kinematics parameters, including amplitude of lateral head displacement (ALH), beat cross frequency (BCF), and curvilinear velocity (VCL), were analyzed using two-way ANOVA to assess the main effects of bag type, storage time, and their interactions. A mixed-effects model was employed to analyze sperm agglutination, incorporating bag type, storage volume, and storage time as fixed effects.
Reproductive outcomes from 1496 sows and 696 gilts were assessed to compare the performance of females inseminated with semen stored in bacteriostatic-coated blister versus conventional blisters. Reproductive parameters, specifically total piglets born and live-born piglets, were analyzed using a Chi-square test.
For all analyses, data are presented as mean ± standard deviation, and statistical significance was set at p ≤ 0.05.

3. Results

3.1. Experiment 1-Bacteriostatic Effects of Bacteriostatic Coating Blister with or Without Antibiotics on Bacterial Contamination

Bacterial contamination across all experimental groups remained generally low, with most measurements below 2 log CFU/mL, indicating effective bacterial control by the bacteriostatic coating blister.
At 120 h of storage, the group utilizing the bacteriostatic coating with antibiotics maintained a bacterial count of 0.365 log CFU/mL, while the group with the coating but without antibiotics registered a higher value of 1.71 log CFU/mL.
During short-term storage (72 h), no significant differences in bacterial growth (CFU/mL, log10) were observed among the bacteriostatic coating blister with antibiotics (0.08 ± 0.27), bacteriostatic coating blister without antibiotics (0.74 ± 0.90), and the Control with antibiotics (0.08 ± 0.19) groups (p > 0.05).
However, during prolonged storage (120 h), bacterial growth was significantly higher in the bacteriostatic coating blister without antibiotic group (1.71 ± 1.18) compared to both the Control with antibiotic (0.195 ± 0.356) and bacteriostatic coating blister with antibiotic (0.365 ± 0.593) groups (p < 0.05). Furthermore, bacterial counts at 120 h were similar between the bacteriostatic coating blister with antibiotic and Control with antibiotic groups (p > 0.05) (Table 1).

3.2. Experiment 1-Bacteriostatic Effects of Bacteriostatic Coating Blister with or Without Antibiotic on Sperm Quality

Table 2 presents the evaluated sperm quality parameters, including total motility, progressive motility, and sperm Kinematics. No statistically significant differences were observed between the experimental groups for any of the analyzed parameters throughout the storage period.

3.3. Experiment 2-Impact of Storage Bag on Swine Semen Quality Under Commercial Conditions

3.3.1. Total and Progressive Motility

Analysis revealed that the type of storage bag significantly influenced total sperm motility (p = 0.015), as presented in Table 3. The bacteriostatic coating blister group exhibited an average total motility of 82.5 ± 10.9%, whereas the conventional (GTB) group showed 81.0 ± 12.6%. There was no significant interaction between bag type and other variables, such as storage duration (p = 0.471), storage volume (p = 0.064), or their combination (p = 0.582). Additionally, no significant interaction was detected between boar breed and bag type for any assessed sperm quality parameters (p > 0.05).
Progressive motility was also significantly higher in the bacteriostatic coating blister group, averaging 65.0 ± 14.0%, compared to 63.1 ± 15.5% for the conventional blister group (p = 0.006). Similar to total motility, the interactions between bag type and storage time (p = 0.08), storage volume (p = 0.28), or the combined effect (p = 0.34), were not statistically significant. Furthermore, no significant decline in these parameters was observed over time (Table 3).

3.3.2. Sperm Kinematics

Significant differences in sperm kinematics parameters were observed between the groups (Table 4). The bacteriostatic coating blister group showed higher mean values for VCL (p = 0.023) and ALH (p < 0.001). In contrast, the conventional blister exhibited higher values for BCF (p < 0.001) and LIN (p = 0.021). No significant overall differences were found for VAP, VSL, and STR.
Beyond these main effects, kinematics analysis revealed a functionally important interaction between storage time and package type for both ALH (p = 0.003) and BCF (p = 0.029). This indicated that the different blister types produced distinct motility profiles over the 168-h period (Table 5). The conventional blister promoted a high-frequency swimming pattern, with BCF values remaining significantly higher than the bacteriostatic group after the initial 24 h. Conversely, the bacteriostatic blister excelled at maintaining a high-amplitude, vigorous movement, particularly over extended storage; by 168 h, its ALH value (8.85 µm) was markedly superior to that of the conventional group (7.98 µm).

3.3.3. Agglutination

The type of storage bag significantly influenced sperm agglutination (p = 0.007). The recorded mean agglutination scores during the storage period were: bacteriostatic coating (40 mL) 2.05 ± 0.972, bacteriostatic coating (80 mL) 2.00 ± 0.989, conventional blister (40 mL) 1.83 ± 0.870, and conventional blister (80 mL) 1.98 ± 1.00. However, no significant interactions were identified between bag type and storage duration (p = 0.616).

3.3.4. In Vivo Trial–Impact of Storage Bag on Reproductive Performance

The reproductive performance of inseminated females did not differ significantly between the groups using blisters with bacteriostatic coating and conventional blisters for either sows (p = 0.66; Table 6) or gilts (p = 0.35; Table 7). The study included a total of 1496 sows (817 inseminated with bacteriostatic-coated doses; 679 with conventional doses) and 696 (353 with bacteriostatic-coated doses; 343 with conventional doses).

4. Discussion

The widespread reliance on stored liquid semen for artificial insemination (AI) in commercial swine production underscores the importance of effective preservation methods. Currently, over 95% of AIs use liquid semen, typically within 24 to 48 h of collection, stored at 15–18 °C to prevent thermal shock to sensitive swine semen [3,7,8,29,30]. This affects the economics of pig production and the rational use of boars.
The reproductive health of a boar directly impacts ejaculate quality, which is also significantly influenced by how semen is collected and processed [31]. Bacteria are naturally present in swine semen from environmental and animal sources, necessitating antibiotics to control their proliferation. These bacteria can vary widely and cause damage to spermatozoa [32,33,34,35]. However, the continuous use of antibiotics has unfortunately led to the emergence of multi-resistant bacteria through epigenetic mechanisms [15,36,37]. This growing concern has driven the search for alternative sperm storage methods that can reduce antibiotic reliance [3,26,38,39].
In industrial pig farming, the development of innovative alternatives to improve semen storage while minimizing antibiotic usage is essential [14]. This study suggests that pig semen can be effectively stored in blisters with bacteriostatic coating without compromising sperm quality or fertility. The hypothesis is that the bacteriostatic agent within BactiBag® does not negatively impact sperm quality, potentially offering an economical alternative as antimicrobial strategies evolve in swine AI. Despite the use of antimicrobials in the field experiment, the study demonstrates that the bacteriostatic coating does not compromise semen quality or reproductive performance in a commercial setting. This technology serves as an alternative for the replacement and/or reduction of antimicrobials in insemination doses. Furthermore, new tests under commercial conditions without the use of antimicrobials could help to consolidate this strategy.
Currently, there is limited evidence demonstrating the efficacy of bacteriostatic strategies for maintaining swine semen doses stored at elevated temperatures (approximately 17 °C) without compromising sperm quality and fertility, particularly in large-scale field insemination contexts. While extenders provide nutrients to sustain sperm longevity for up to seven days, they also inadvertently create favorable conditions for the growth of both natural and antimicrobial-resistant contaminant microorganisms, making prolonged viability challenging due to bacterial proliferation [10,40].
Bacterial contamination, or bacteriospermia, negatively impacts reproductive performance, with the severity directly proportional to bacterial concentration [29]. Bacteria affect sperm quality through various mechanisms, including agglutination, reduced motility, altered acrosome function, and loss of plasma membrane integrity [40]. Recent reviews further highlight detrimental effects of bacterial growth on sperm, such as increased apoptosis, reduced mitochondrial membrane potential, and elevated reactive oxygen species (ROS) levels, all exacerbating damage to sperm cells [10].
For swine semen doses to be commercially viable, they must meet strict criteria: sperm motility must remain between 45% and 70% until expiration, and bacterial contamination must not exceed 1000 CFU/mL (equivalent to 2–3 log) [27]. The peak of bacterial growth in semen doses typically occurs after 48 h of storage, marking a critical point for maintaining quality. Although antibiotics in extenders are effective, they are not a long-term solution [41,42]. Consequently, the market is increasingly trending towards alternatives that allow for the reduction and rational use of antibiotics in semen doses.
The physical removal of bacteria through methods like colloid centrifugation has emerged as a promising alternative to antibiotics. This technique, proven effective for swine semen on a laboratory scale, physically separates spermatozoa from bacteria without impairing sperm quality [21,22,23,24,34]. Nevertheless, its transition to large-scale use in AI centers is hindered by practical and financial hurdles. Colloid centrifugation introduces an additional, costly step—requiring specialized training, equipment, and materials—that can elevate the final price by about US$2 per dose [21].
This economic barrier underscores the need for more accessible innovations. The BactiBag system, for example, presents a starkly different cost-benefit profile. While it increases the packaging cost by 15–20% relative to a standard GTB BAG blister, this translates to a minimal absolute increase of approximately €0.02 to €0.03 per dose. In the Brazilian context, this means the packaging’s share of the total production cost would shift only marginally from 9.55% to 11.05%. Therefore, evaluating sustainable technologies in swine semen production requires a crucial balance between proven biological efficacy and overall economic feasibility.
Blisters with bacteriostatic coating emerge as a promising solution in this context. These blisters are engineered with a weldable thermoplastic layer that contains a phenolic antimicrobial compound (5-Chloro-2-(2,4-dichlorophenoxy)phenol; CAS N° 3380-34-5) [43,44]. This compound functions as a bacteriostatic agent, effectively controlling bacterial growth while ensuring minimal release into the semen. The product acts by direct contact on the bag’s surface, a design that minimizes sperm exposure to the antimicrobial agent, thereby preserving their motility and viability [43]. Unlike studies where the substance directly contacted sperm, leading to adverse effects like loss of motility and mitochondrial depolarization [45], BactiBag® employs a physical barrier. This approach effectively manages the antimicrobial effect without compromising sperm quality or cellular integrity.
Although the present study did not independently validate the antimicrobial efficacy of the blister coating, the technology’s underlying principles and specifications are detailed in its patent documentation [43]. The bacteriostatic agent is incorporated into the inner polyethylene layer of the blister. The patent specifies that the concentration of the phenolic antimicrobial compound ranges from 1 mg/m2 to 40 mg/m2. Furthermore, the patent includes in vitro studies demonstrating that this concentration is effective in controlling bacterial proliferation in antibiotic-free diluted boar semen, maintaining bacteriostatic conditions when compared to conventional packaging. Therefore, our study’s hypothesis was based on this pre-existing and validated technological foundation, and our focus was to assess its impact on sperm quality and reproductive performance under large-scale commercial conditions.
In this study, the bacteriostatic coating blister demonstrated effective bacterial control, even without antibiotics, keeping bacterial counts below 2 log10. This result is comparable to the efficacy achieved with gentamicin in extenders in both control and bacteriostatic-coated blister groups with antibiotics, aligning with findings from other research [41,42]. Furthermore, sperm characteristics remained consistent in both the control and bacteriostatic-coated blister groups, regardless of antibiotic presence, throughout the entire storage period. These findings suggest that the bacteriostatic coating blister, irrespective of antibiotic use, did not compromise the sperm quality of the evaluated doses compared to the control. The stability of these parameters indicates the suitability of the BactiBag® to preserve sperm functionality under various experimental conditions. Additionally, these findings underscore the safety and efficacy of using the bacteriostatic plastic blister for swine semen storage, highlighting its ability to balance bacterial control with the preservation of sperm quality.
The seminal quality of pigs is influenced by various factors that can compromise reproductive potential [46]. Therefore, new methods aimed at substituting or reducing antibiotic use must effectively control bacterial proliferation during storage while preserving sperm parameters essential for efficient fertilization. Moreover, innovative solutions that maintain satisfactory reproductive outcomes will significantly contribute to minimizing antimicrobial use in pig farming, thereby mitigating the spread of bacterial resistance.
To address these critical requirements, this study evaluated the bacteriostatic coating blister’s efficacy in preserving swine sperm quality for up to seven days (168 h) and its ability to mitigate bacterial growth. The analyses included total motility, progressive motility, sperm Kinematics, and sperm agglutination, all performed according to industry standards [3]. The results conclusively showed that BactiBag® maintained total motility above 75% throughout the storage period, alongside significant improvements in progressive motility.
Previous research highlights that sperm motility below 60% can compromise fertility, leading to reduced in vitro sperm penetration rates, farrowing rates, and smaller litter sizes [6]. Crucially, both groups maintained total motility above 75% and progressive motility above 50% across all assessed storage times (24, 72, 120, and 168 h). Among various sperm parameters, progressive motility is particularly important as it indicates cell metabolism and membrane integrity, directly influencing field fertility and farrowing rates in sows [29,47]. The use of CASA systems has enabled objective and detailed evaluation of sperm kinematics parameters, which are vital for predicting fertility accurately in commercial settings [6].
Sperm kinematics assessment, especially through CASA, serves as an important indicator of semen quality and fertility potential across species like cattle and swine. Analyzing kinematics parameters provides an objective understanding of sperm movement and has shown a connection to fertility outcomes, though this link’s precision is still under investigation [48,49]. Our results demonstrated that the type of packaging distinctly influenced sperm kinematics patterns, which are critical indicators of fertility potential [47]. The fact that the bacteriostatic blister promoted a more vigorous motility profile (higher ALH and VCL) supports our hypothesis that its primary benefit comes from superior bacterial control. We believe that by limiting bacterial proliferation and the subsequent release of endotoxins, the structural integrity and energetic capacity of spermatozoa were better preserved. This sustained energy enabled a high-amplitude swimming pattern. While this did not translate to higher final fertility in this specific study, it is a strong indicator of the higher quality and viability of the semen dose during extended storage. These findings are further reinforced by the consistent maintenance of higher total and progressive motility in the bacteriostatic blister group under field conditions, confirming its protective effect on sperm function.
This study also included sperm agglutination analysis as an in vitro assay, which unexpectedly showed a higher incidence of agglutination in samples stored in bacteriostatic-coated blisters. Traditionally, agglutination in swine semen has been linked to bacteriospermia and reduced sperm motility [40,50]. However, some studies have reported sperm agglutination without identifying contaminating microorganisms [11]. Additionally, Delgado-Bermúdez et al. [51] found no direct relationship between sperm motility alterations caused by Proteus vulgaris and sperm agglutination, suggesting the involvement of other factors in this phenomenon. Notably, the mean agglutination at all evaluated time points remained below 30% throughout the 168 h. This suggests that, in this storage context, controlled sperm agglutination does not compromise reproductive functionality.
In the present study, the findings indicate that the observed sperm agglutination did not adversely affect overall semen quality, specifically total and progressive motility. This suggests that agglutination occurrences here might not be linked to bacteriospermia, possibly arising from alternative mechanisms. Given the important role of sperm-sperm interaction in sperm selection and capacitation for fertilization [52], agglutination could potentially be part of the natural reactivation process of cells after storage. In both evaluated groups, the agglutination rate remained below 30% in insemination doses, and key sperm parameters stayed within satisfactory limits for AI. Our findings suggest that controlled sperm agglutination, in this storage context, does not compromise reproductive functionality.
These findings highlight the need for further research to clarify the biological mechanisms underlying sperm agglutination across various storage methods. Future studies could provide valuable insights into how different storage conditions impact this phenomenon, ultimately contributing to advancements in swine semen preservation technologies.
This study primarily focused on sperm quality parameters directly associated with field fertility outcomes, specifically kinematics and agglutination. However, a more comprehensive evaluation of sperm physiology was not performed. Analyzing attributes such as plasma and acrosomal membrane integrity, mitochondrial membrane potential, and chromatin integrity would offer a more complete and mechanistic understanding of the bacteriostatic blister’s effects, particularly during prolonged storage periods.
While CASA and the assessment of conception and birth rates provide objective and predictive data highly relevant to commercial settings, they represent only one facet of complex sperm viability. Consequently, the absence of a more detailed physiological analysis is acknowledged as a methodological limitation of this study.
Future research should consider incorporating advanced techniques, such as flow cytometry, to investigate these subcellular parameters. Such an in-depth approach would complement the robust reproductive performance results presented herein, simultaneously validating the technology’s safety and impact at a more detailed physiological level.
Our data further reinforce that sperm stored in bacteriostatic-coated blisters, utilizing extenders for prolonged storage (16–18 °C), maintain adequate levels of total and progressive motility to satisfy AI demands under commercial conditions. This approach not only enhances operational flexibility for semen dose utilization but also aligns with sustainable reproductive strategies within the framework of the One Health concept [17].
Previous investigations have demonstrated the potential of BactiBag® in controlling bacterial growth and preserving semen quality up to three days [44]. This study extends that understanding, showing the efficacy of BactiBag® in maintaining sperm viability for up to seven days. Importantly, this blister represents a significant advancement in swine reproduction technology by contributing to the reduction or potential elimination of antimicrobials in semen extenders [3,53]. Although the present study did not conduct fully antibiotic-free experiments, it provides a valuable foundation for future research aimed at assessing fertility in large-scale inseminations under real operational conditions.
The interpretation of the in vitro results consistently focused on their applicability to in vivo AI performance within a productive context. Given the current reproductive efficiency in the swine industry, with farrowing rates frequently exceeding 90% [7,54,55], our trials were specifically designed to address economically relevant issues, particularly those stemming from compromised AI dose quality [11].
The effectiveness of the bacteriostatic coating blister was strongly corroborated by a large-scale field trial, demonstrating fertility results comparable to conventional storage practices. This research also highlighted the viability of using these blisters across diverse productive categories, including multiparous sows and nulliparous females (gilts). Despite the inherent challenges of conducting experiments within commercial systems, the proposed strategy successfully enabled the standardization of experimental conditions necessary for robust comparison among pigs [56].
While this technology aligns with sustainable practices by reducing the reliance on antibiotics—a key goal within the One Health framework—it is imperative to address the environmental and regulatory aspects of using the bacteriostatic agent. The claim of sustainability is nuanced, as substituting antibiotics in the extender with a biocidal agent in the packaging material shifts the environmental challenge from the use phase to the disposal phase.
Triclosan is recognized for its environmental toxicity, particularly to aquatic ecosystems. Consequently, the post-use blister cannot be managed as conventional plastic waste. According to environmental and sanitary regulations in many jurisdictions, such as Brazil (e.g., ANVISA RDC 222/2018 and ABNT NBR 10004) and the European Union (under the European List of Waste-LoW), a product containing an active antimicrobial agent is classified as hazardous waste. This approach contrasts with the regulatory landscape in the United States, where, under the Resource Conservation and Recovery Act (RCRA), Triclosan is not listed as a hazardous waste. As a result, unless the material fails a specific characteristic test (e.g., for toxicity, ignitability), the used blister can often be managed as non-hazardous solid waste.
In regions with stricter regulations, the hazardous waste classification mandates a strict disposal protocol to prevent environmental contamination. The required post-use management includes segregating the used blisters at the point of generation in designated containers for chemical waste, which must be clearly labeled with the appropriate hazard symbol. These materials must then be collected by a licensed waste management company for appropriate treatment and final disposal. The recommended treatment method is incineration in a specialized facility, which ensures the destruction of the hazardous chemical compound. The final destination for any resulting ash or the untreated product is a Class I hazardous waste landfill, engineered to contain toxic substances and prevent soil and groundwater contamination. Therefore, while the bacteriostatic blister is a significant advancement in combating antimicrobial resistance, its overall sustainability is contingent upon the implementation of these rigorous and legally required waste management practices, the stringency of which varies significantly between countries.
Crucially, the main reproductive parameters evaluated—farrowing rate, total number of piglets born (TNB), number of live-born piglets, and total litter weight—were not negatively impacted by using either bacteriostatic-coated or conventional blisters in both sows and gilts [7]. Reproductive outcomes, including the total number of piglets born and the number of live-born piglets, aligned with expected performance levels in high-prolificacy females. These findings definitively demonstrate that the blister with a bacteriostatic coating is a viable alternative or complement to existing antimicrobial methods for controlling bacterial contamination in semen. This innovative solution holds significant potential to enhance the reproductive performance of females and contribute directly to the advancement of commercial swine production.

5. Conclusions

This study conclusively demonstrates that swine spermatozoa can be effectively stored in blisters with a bacteriostatic coating without compromising fertility. The research confirmed the effectiveness of these blisters in preserving sperm viability for up to seven days (168 h). Although the large-scale commercial farm trial (Experiment 2) did not directly assess the bacteriostatic properties of the blister, the in vitro results from Experiment 1 confirmed its efficacy in controlling bacterial growth. This validation showcases its ability to protect semen quality and ensure consistent reproductive performance.
This technology offers enhanced flexibility in swine semen storage by extending viability while maintaining all essential sperm parameters for artificial insemination. Crucially, the adoption of bacteriostatic-coated blisters aligns with more sustainable reproductive practices and directly addresses the increasing demand for reduced antibiotic use in livestock. This represents a significant advancement in commercial swine production.

Author Contributions

Conceptualization, P.N.J.-N. and R.Z.; methodology, J.d.C., P.N.J.-N., J.V.B. and R.Z.; validation, J.d.C., J.V.B. and R.Z.; formal analysis, J.d.C. and R.P.; investigation, J.d.C., É.L.M., M.G.D.d.O. and G.F.; writing—original draft preparation, J.d.C., P.N.J.-N., R.P. and R.Z.; writing—review and editing, J.d.C., P.N.J.-N., J.V.B., R.P. and R.Z.; visualization, J.d.C., P.N.J.-N., R.P. and R.Z.; supervision, P.N.J.-N. and R.Z.; project administration, R.Z.; funding acquisition, R.Z. All authors have read and agreed to the published version of the manuscript.

Funding

CNPq MAI DAI Scholarship-Process: 157824/2021-7, and CNPq Productivity in Research Scholarship 2–Process: 301636/2022-2.

Institutional Review Board Statement

This study received approval from the Animal Use Ethics Committee of the University of Passo Fundo, Brazil (CEUA-UPF), under protocol No. 005/2023.

Data Availability Statement

The datasets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.

Acknowledgments

The authors acknowledge Nutribrás Alimentos and its contribution to this work.

Conflicts of Interest

J.d.C. and R.Z. report that financial support was provided by the National Council for Scientific and Technological Development. P.N.J.-N. reports a relationship with IMV do Brasil that includes employment. R.Z. reports a relationship with IMV do Brasil that includes consulting or advisory roles. To avoid any ethical concerns about the results, P.N.J.-N. and R.Z. did not participate in the experimental activities or data collection. The other authors declare no known competing financial interests or personal relationships that could have influenced the work reported in this paper.

Abbreviations

The following abbreviations are used in this manuscript:
AIArtificial Insemination
ALHAmplitude of Lateral Head Displacement
BCFBeat Cross Frequency
BTSBeltsville Thawing Solution
CASAComputer-Assisted Semen Analysis
CFU/mLColony-Forming Units per milliliter
EDTAEthylenediaminetetraacetic Acid Disodium Salt Dihydrate
LINLinearity
PBSPhosphate Buffered Saline
PCAIPost-Cervical Artificial Insemination
ROSReactive Oxygen Species
STRStraightness
TNBTotal Number of Piglets Born
VAPAverage Path Velocity
VCLCurvilinear Velocity
VSLStraight-Line Velocity

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Table 1. Bacterial Contamination in Semen Storage (n = 11 ejaculates). This table shows the least square means ± SD for bacterial contamination in semen stored in different blisters over time.
Table 1. Bacterial Contamination in Semen Storage (n = 11 ejaculates). This table shows the least square means ± SD for bacterial contamination in semen stored in different blisters over time.
Storage Time
(Hours)		Group
BB + A (n = 22)BB − A (n = 22)CB + A (n = 22)CB − A (n = 22)
Bacterial Proliferation
(log)		720.08 ± 0.27 a0.74 ± 0.90 ab0.08 ± 0.19 a1.64 ± 1.82 b
1200.365 ± 0.59 a1.71 ± 1.18 b0.195 ± 0.35 a2.04 ± 1.67 b
BB + A: Bacteriostatic Coating Blister with Antibiotic; BB − A: Bacteriostatic Coating Blister without Antibiotic; CB + A: Conventional Blister with Antibiotic; CB − A: Conventional Blister without Antibiotic. Values with different letters on the same line differed from each other (p < 0.05).
Table 2. Boar (n = 5) Sperm Characteristics in Semen Storage (n = 10 inseminate dose per treatment). The table presents the least square means ± SD for various sperm parameters at different storage times.
Table 2. Boar (n = 5) Sperm Characteristics in Semen Storage (n = 10 inseminate dose per treatment). The table presents the least square means ± SD for various sperm parameters at different storage times.
Sperm ParametersStorage Time
(Hours)		Groupp Value
BB + A (n = 10)BB − A (n = 10)CB + A (n = 10)CB − A (n = 10)
Total Motility, %7287.8 ± 11.985.8 ± 11.681.7 ± 18.483.1 ± 9.3NS
12073.9 ± 21.481.3 ± 13.873.7 ± 17.179.1 ± 15.1NS
Progressive Motility, %7242.4 ± 18.535.7 ± 14.231.0 ± 7.027.6 ± 7.1NS
12035.4 ± 21.724 ± 16.123.3 ± 15.921.7 ± 6.9NS
ALH, µm726.87 ± 0.806.75 ± 0.806.03 ± 0.606.27 ± 1.18NS
1206.88 ± 0.675.92 ± 2.165.39 ± 1.074.89 ± 0.87NS
BCF, Hz7234.2 ± 2.8434.7 ± 3.2134.0 ± 2.7235.0 ± 4.34NS
12038.2 ± 5.8534.9 ± 4.0437.3 ± 6.2840.6 ± 7.69NS
VCL, µm/s72128.0 ± 29.0121.0 ± 25.0110.0 ± 18.6108.0 ± 25.6NS
120123.0 ± 27.6105.0 ± 49.391.5 ± 27.087.0 ± 15.3NS
VAP, µm/s7261.4 ± 15.557.4 ± 11.853.3 ± 6.349.5 ± 10.8NS
12057.3 ± 17.148.8 ± 23.442.9 ± 14.440.5 ± 1.3NS
VSL, µm/s7240.5 ± 11.737.2 ± 9.035.6 ± 6.330.8 ± 4.7NS
12036.5 ± 11.030.9 ± 11.627.8 ± 10.227.9 ± 4.6NS
STR, VSL/VAP %7267.0 ± 7.066.0 ± 6.369.4 ± 8.564.8 ± 11.2NS
12063.1 ± 3.667.7 ± 12.368.8 ± 3.369.1 ± 11.7NS
LIN, %7235.2 ± 6.734.2 ± 6.537.7 ± 10.833.5 ± 11.7NS
12031.6 ± 4.236.2 ± 13.135.7 ± 5.537.1 ± 13.0NS
BB + A: Bacteriostatic Coating Blister with Antibiotic; BB − A: Bacteriostatic Coating Blister without Antibiotic; CB + A: Conventional Blister with Antibiotic; CB − A: Conventional Blister without Antibiotic.
Table 3. Boar Sperm Characteristics in Semen Storage: Bacteriostatic Coating Blister vs. Conventional Blister. The table displays least square means ± SD for sperm characteristics at various storage points.
Table 3. Boar Sperm Characteristics in Semen Storage: Bacteriostatic Coating Blister vs. Conventional Blister. The table displays least square means ± SD for sperm characteristics at various storage points.
Storage PointGroupnTotal Motility, %p ValueProgressive Motility, %p Value
D0BB13787.5 ± 9.05*75.8 ± 11.2*
CB13488.7 ± 6.77*77.0 ± 9.93*
D24BB13681.5 ± 10.0NS63.0 ± 13.4NS
CB13280.0 ± 10.8NS62.0 ± 14.5NS
D72BB13581.2 ± 11.4NS60.3 ± 14.1NS
CB13379.3 ± 12.8NS58.5 ± 15.7NS
D120BB13782.6 ± 10.2NS64.3 ± 12.2NS
CB13479.4 ± 12.8NS59.4 ± 14.7NS
D168BB13579.7 ± 12.1NS61.6 ± 13.4NS
CB13177.6 ± 15.2NS58.1 ± 15.8NS
BB: Bacteriostatic Coating Blister; CB: Conventional Blister; * Not applicable for baseline/fresh semen; NS = not significant (p > 0.05) for comparisons between groups (bacteriostatic blister vs. conventional).
Table 4. Summary of Sperm Kinematics Parameters (Overall Group Effect).
Table 4. Summary of Sperm Kinematics Parameters (Overall Group Effect).
Sperm Kinematics ParameterBacteriostatic BlisterConventional Blisterp-Value
ALH (µm)8.27 ± 1.527.93 ± 1.57<0.001
VCL (µm/s)191.7 ± 38.0187.3 ± 40.60.023
BCF (Hz)36.1 ± 2.6536.6 ± 2.76<0.001
LIN (%)33.1 ± 7.9034.1 ± 8.740.021
Table 5. Interaction Effects between Storage Bag and Time on ALH and BCF Parameters.
Table 5. Interaction Effects between Storage Bag and Time on ALH and BCF Parameters.
Storage Time PointsALHBCF
CBBBCBBB
D08.48 ± 2.07 ab8.33 ± 2.12 abc34.7 ± 3.16 d34.7 ± 3.12 d
D247.56 ± 1.49 d7.93 ± 1.30 cd37.7 ± 2.34 a36.8 ± 2.60 b
D727.80 ± 1.26 cd8.26 ± 1.21 abc37.9 ± 2.31 a36.9 ± 2.47 b
D1207.83 ± 1.34 cd8.22 ± 1.33 bc37.1 ± 2.39 ab36.3 ± 2.23 c
D1687.98 ± 1.44 c8.85 ± 1.14 a36.4 ± 2.36 c35.2 ± 1.95 d
BB: Bacteriostatic Coating Blister; CB: Conventional Blister. Least square means ± standard deviation are shown. For each parameter (ALH and BCF sepahrately), means followed by different superscript letters are significantly different (p < 0.05) according to Tukey’s HSD test.
Table 6. Reproductive Performance of Inseminated Sows.
Table 6. Reproductive Performance of Inseminated Sows.
Performance ParametersConventional BlisterBacteriostatic Blister
Sows (n)679817
Pregnancy rate (day 30)93.5% (635/679)93% (760/817)
Farrowing rate92.7% (630/679)92.4% (755/817)
Average Number of Piglets Born per Litter16.1 ± 3.8816.0 ± 4.13
Average Number of Live-Born Piglets per Litter14.5 ± 3.7714.4 ± 3.85
Average Total Litter Weight (kg)19.6 ± 5.1219.2 ± 5.08
Values in parentheses represent the count of animals for pregnancy and farrowing rates. The averages for born and live-born piglets are presented as mean ± standard deviation.
Table 7. Reproductive Performance of Inseminated Gilts.
Table 7. Reproductive Performance of Inseminated Gilts.
Performance ParametersConventional BlisterBacteriostatic Blister
Gilts (n)343353
Pregnancy rate (day 30)95% (326/343)99.1% (350/353)
Farrowing rate93.2% (320/343)94.6% (334/353)
Average Number of Piglets Born per Litter14.7 ± 3.5314.9 ± 4.07
Average Number of Live-Born Piglets per Litter13.6 ± 3.3113.7 ± 3.58
Average Total Litter Weight (kg)18.1 ± 4.2618.1 ± 4.71
Values in parentheses represent the count of animals for pregnancy and farrowing rates. The averages for born and live-born piglets are presented as mean ± standard deviation.
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MDPI and ACS Style

Camargo, J.d.; Jorge-Neto, P.N.; Madruga, É.L.; Oliveira, M.G.D.d.; Fruhling, G.; Braga, J.V.; Poletto, R.; Zanella, R. Innovation on Swine Semen Storage: Bacteriostatic Coating vs. Conventional Blister in Commercial Swine Semen Production. AgriEngineering 2025, 7, 338. https://doi.org/10.3390/agriengineering7100338

AMA Style

Camargo Jd, Jorge-Neto PN, Madruga ÉL, Oliveira MGDd, Fruhling G, Braga JV, Poletto R, Zanella R. Innovation on Swine Semen Storage: Bacteriostatic Coating vs. Conventional Blister in Commercial Swine Semen Production. AgriEngineering. 2025; 7(10):338. https://doi.org/10.3390/agriengineering7100338

Chicago/Turabian Style

Camargo, Janine de, Pedro Nacib Jorge-Neto, Érika Lopes Madruga, Maria Gessica Daniel de Oliveira, Gilson Fruhling, José Victor Braga, Rosangela Poletto, and Ricardo Zanella. 2025. "Innovation on Swine Semen Storage: Bacteriostatic Coating vs. Conventional Blister in Commercial Swine Semen Production" AgriEngineering 7, no. 10: 338. https://doi.org/10.3390/agriengineering7100338

APA Style

Camargo, J. d., Jorge-Neto, P. N., Madruga, É. L., Oliveira, M. G. D. d., Fruhling, G., Braga, J. V., Poletto, R., & Zanella, R. (2025). Innovation on Swine Semen Storage: Bacteriostatic Coating vs. Conventional Blister in Commercial Swine Semen Production. AgriEngineering, 7(10), 338. https://doi.org/10.3390/agriengineering7100338

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