Bacteriospermia among smallholder artificial insemination boars in the Philippines and potential associated factors

Objective:To determine the prevalence of bacteriospermia,the bacterial load,and the potential factors associated with bacterial contamination in boar semen collected by local smallholder artificial insemination operators.Methods:Fifteen individual raw semen samples were collected from locally available artificial insemination boars owned by different smallholder boar operators within the 5th district of Leyte,Philippines and were subjected to standard bacteriological culture and identification,including a survey of potentially associated factors.Prevalence and bacterial count were determined accordingly,while boar characteristics and collection practices were clustered following agglomerative hierarchical clustering technique.Results:One hundred percent contamination with a bacterial count of(2.01±0.38)×10^(3) CFU/mL was observed.At least 73.33%of the samples were positive for Bacillus spp.,while other identified isolates included Enterobacter spp.,Staphylococcus spp.,E.coli,Pseudomonas spp.,Citrobacter spp.,and Klebsiella spp.Conclusions:Despite the high prevalence of bacteriospermia,the bacterial count is low.Nevertheless,on-farm practices on boar health and management,semen collection,and sanitation as well as the enhancement of basic protocols to control contamination should be conscientiously considered in smallholder artificial insemination operation.

Serologic and molecular survey for major viral pathogens in grazing hybrid wild boars in Northeast China

Hybrid wild boar husbandry is an important component of livestock production in Northeast China.However,the current disease situation of these animals is largely unknown due to a lack of disease surveillance.The present study was conducted to determine the prevalence of several important viral diseases in the hybrid wild boar population of Northeast China.Between September 2015 to December 2016,169 blood and 61 tissue samples were collected from apparently healthy hybrid wild boars from farms in Jilin,Inner Mongolia and Heilongjiang provinces.ELISA detected serum antibodies against classical swine fever virus(CSFV),porcine reproductive and respiratory syndrome virus(PRRSV),pseudorabies virus(PRV),porcine circovirus type 2(PCV2)and Japanese encephalitis virus(JEV),but not against African swine fever virus(ASFV),with PCV2 having the highest seropositive rate(87.2–100%in different farms).RT-PCR or PCR performed on the processed samples detected only PCV2,with 33.1%(56/169)of blood samples and 32.8%(20/61)of spleen samples being positive,respectively,indicating widespread PCV2 infection in hybrid wild boars.Phylogenetic analysis of 15 PCV2 ORF2 sequences showed that they belong to genotypes PCV2a,PCV2b and PCV2d,with nucleotide and deduced amino acid homologies of 88.5–100%and 88.1–100%,respectively.

Analysis of the lifetime and culling reasons for AI boars

Background: The aim of the study was to analyze the lifetime and culling reasons for boars used in insemination centers(AI centers).Methods: The data collected from 355 culled boars from 1998 to 2013 included: age at start of semen collection,boar herd life, culling reason, daily gain and lean meat content, and number of ejaculates not meeting sales requirements after dilution. Culling reasons were divided into 7 groups: low semen value(LSV), low or lack of libido(LL), leg problems(LP), infectious diseases(ID), old age(OA), reduced demand for semen from the given boar(RD),and others(OT).Results: The most common culling reasons for boars were LSV(23.7%) and RD(22.5%). It was observed that the lowest daily gains were noted in boars culled due to OA. Boars culled due to OA and RD were maintained in production for the longest time(over 1000 d), for LSV and ID retention was about 700 d, and due to LL below 400 d. The survival probability was over 0.9 until 1.5 yr, and just over 0.2 until 4 yr. The highest relative frequency was observed in the 36 ^(th) and 42 ^(nd) mo of life(over 16%). Hazard risk analysis revealed a more than 10 times higher risk of culling in the case of LL, ID or OT, in comparison to OA.Conclusions: The results can be used as a direct point of reference for the identification of emerging problems in AI boar exploitation and the development of an appropriate culling policy in AI centers.

Semen characteristics of the three genetic types of boars reared in Benin

Objective:To characterize the semen of three genetic types of boars(local,improved and Large White)reared in Benin.Methods:Semen of local,improved and Large White boars reared in Benin were collected using the gloved hand method and analyzed to determine volume,pH,concentration,mobility,motility,and morphology.The effect of the genetic type of boar on semen characteristics was aslo studied.Results:Duration of ejaculation and semen volume of Large White boar were significantly higher than those of local and improved boars(P<0.05).The semen of improved boars had a higher motility score than that of Large White and local boars(P<0.001).The semen of local boars was more concentrated in the spermatozoa than that of improved and Large White boars(P<0.05).The proportion of spermatozoa of improved boars with normal morphology(93.6%)was significantly higher than that of local(82.2%)and Large White boars(81.6%)(P<0.001).The proportion of spermatozoa with folded tails in the semen of Large White boars(9.2%)was significantly higher than that observed in improved(1.8%)and local(5.0%)boars(P<0.001).The proportion of spermatozoa with proximal cytoplasmic droplets in semen of improved boars(2.7%)was significantly lower than that in Large White(6.8%)and local(9.7%)boars(P<0.001).The local(1.5%)and Large White boars(1.1%)showed more spermatozoa with distal cytoplasmic droplets in their semen compared to the improved boars(0.4%).Conclusions:The semen characteristics of pigs reared in Benin vary from one genetic type to another.Each genetic type has a strong point.The Large White boar produces larger semen,the local boar produces more concentrated semen and the improved boar produces spermatozoa that are morphologically better.The semen of these three genetic types can be used in artificial insemination but the improved boar's semen is more recommended.

Effects of Selenium-Enriched Probiotics on Semen Quality of Stock Boars

Selenium-enriched probiotics were supplemented into feedstuff of Landrace stock boars at a concentration of 0.3 mg/kg （concentration of selenium）, and the semen quality of the stock boars were observed. The results showed that the semen motility, rate of intact acrosome, and activity of glutathione peroxidase were higher in the experimental boars than in the control, but rate of sperm deformity was lower in the experimental boars than in the control. Therefore, the selenium-enriched probiotics greatly improve semen quality of stock boars.

Evaluation of the Nutritional Supplement VIUSID Vet Powder on the Productive Behaviour of Sows and Boars

The objective of this research was to evaluate the effect of administering VIUSID vet powder on the productive behaviour of sows and the quality of semen of boars. Two experiments were conducted on farms in the province of Sancti Spiritus, in the central region of Cuba. One was with sows, starting 21 days before parturition until weaning, and another was with boars in directed mounting. Each sow received 10 grams of VIUSID vet powder everyday, mixed with food, from 21 days before the probable date of farrowing and until weaning （33-day lactation period）. In the boars, the treatment consisted of administering 10 grams of VIUSID vet powder with feed, once a day for 6 weeks （week 1 to 6）; from week 7 onwards, the supplement was suspended. Results showed that, supplying VIUSID vet powder significantly improved the principal post-partum productivity indicators and significantly reduced diarrhoea and administration to boars in production for 6 weeks, which has improved spermatic motility and does not affect the other variables representative of the quality of the semen.

Effect of Stabilized Fish Oil Source on Sperm Quality and Production of Boars

Research findings for supplementing boar stud diets with fish oils are inconsistent. This study was designed to address three possible causes of performance variation of boars to fish oil supplementation: stability of the fatty acid source, level of inclusion and breed of boars tested. Three groups of 87 boars each, from two genetic lines (PIC 337 and PIC 800), were assigned to treatment based on age, mean sperm production (previous 12 weeks), and body condition score. All boars received a corn-soybean meal diet with a commercial fish oil supplement providing 1.83 g/boar/day of docosahexaenoic acid (DHA) as a preconditioning diet. On 10-Aug., 2020, the DHA source was changed to a stabilized starch imbedded source of refined fish oil (Salmate®), providing 1.83 g/b/d for the test diet. Two additional levels providing 2.38 and 2.94 g/b/d of DHA were fed for a 9 week pretreatment period and during the test period. Salmate® fed at 2.38 g/b/d of DHA resulted in a reduction in the number of rejected ejaculates (P < 0.045) by 7.5% and 6.4% compared to the lowest and highest inclusion rates, respectively. There were no treatments by genetic line interactions. A retrospective study of semen production and quality of 77 boars on the Salmate® diet containing 1.83 g/b/d DHA was done to compare to the original source of DHA at the same inclusion level. There were no differences in semen quality parameters between the 2 lipid sources. Ejaculate volume increased from 177.9 ml to 233.4 ml (P < 0.001) and total sperm cells per ejaculate increased from 69.7 × 109 to 82.0 × 109 (P < 0.001) due to substitution of Salmate®. Adding Salmate® at 2.38 g/b/d resulted in a lower number of rejected ejaculates per boar by 7.5% and 6.4% vs. 1.83 and 2.94 g/b/d, respectively, and boars fed Salmate® at 1.83 g/b/d produced 17% more doses than the competing product.

Conflict between wild boars(Sus scrofa) and farmers:distribution, impacts,and suggestions for management of wild boars in the Three Gorges Reservoir Area

Economic losses caused by wild boars(Sus scrofa)continue to increase in the Three Gorges Reservoir Area.We selected 206 farmers in Wuqiaohe Basin as survey respondents using random stratified sampling.The distribution and impacts of wild boars were examined using a questionnaire survey and spatial analysis.The results showed that:(1)The density of farmer settlements is low and most peasants’households are located on the forest edge.Due to large-scale labor migration,a high proportion of farmland(25.85%)has been abandoned and an even higher proportion of farmland(40.48%)has been returned to forest.(2)The probability of wild boar damage to peasant households was 67.96%.Yields of corn and sweet potato decreased by 24.87%and 28.24%,respectively,with a total economic loss of approximately 2,590,100 RMB/yr in the Wuqiaohe basin.Sixty-five percent of the affected cultivatedlands were located within approximately 150 m of the forest edges.(3)The wild boar damage coefficient was significantly and negatively correlated with the distance between the peasants’household and the forest edge,the density of the settlements,and the area of cultivated land.The wild boar damage coefficient shows a significant positive correlation with the area of farmland returned to forest.(4)The damage activity caused by wild boars began to occur7-8 years ago,and the destruction has been more frequently and seriously observed in the last 4-5 years.(5)We make suggestions for wild boar control and management measures including developing a compensation policy.The compensation standard is related to the land area returning from farmland to forests and the actual cultivated land area.

Effects of pituitary-specific overexpression of FSHα/β on reproductive traits in transgenic boars

Background： Follicle-stimulating hormone（FSH） is a gonadotropin synthesized and secreted by the pituitary gland.FSH stimulates follicle development and maturation in females. It also plays an important role in spermatogenesis in males, including humans and mice. However, the effects of FSH on male pigs are largely unknown. In this study,we generated transgenic pigs to investigate the effects of FSHα/β overexpression on reproductive traits in boars.Results： After five transgenic F0 founders were crossed with wide-type pigs, 193 F_1 animals were obtained. Of these,96 were confirmed as transgenic. FSHα and FSHβ m RNAs were detected only in pituitary tissue. Transgenic boars exhibited significantly higher levels of FSHα and FSHβ m RNA, serum FSH, and serum testosterone, compared to ful-sib non-transgenic boars. Significant increases in testis weight, vas deferens diameter, seminiferous tubule diameter,and the number of Leydig cel s were observed, suggesting that the exogenous FSHα/β affects reproductive traits. Final y,transgenic and non-transgenic boars had similar growth performance and biochemical profiles.Conclusions： Pituitary-specific overexpression of FSHα/β genes is likely to impact reproductive traits positively, as indicated by enhancements in serum testosterone level, testis weight, the development of vas deferens, seminiferous tubules, and Leydig cells in transgenic boars. A high level of serum FSH induces secretion of serum testosterone,possibly by boosting the number of Leydig cells, which presumably increases the libido and the frequency of sexual activity in transgenic boars. Our study provides a preliminary foundation for the genetic improvement of reproductive traits in male pigs.

Genetics Research and Advance on Development and Utilization of Wild Boars

Wild boar is one of the most important beast resources. It plays an important role in the maintenance of biological diversity. The genetic resources of wild boar can not only protect the genetic resources, but also improve the formation of new breeds in pigs. This paper summarized the advance on the main biological characteristics of wild boars, evolutionary origin between wild boars and domesticated pigs, and development and utilization of wild boars aimed to provide further insight into wild boar＇s genetic research and its resource protection.

Relationship between Serum L-Carnitine Levels and Sperm Parameters in Boars

This study evaluated the relationship between serum L-carnitine level and sperm parameters in young boars. Serum L-carnitine and semen characteristics were determined for 61 young Duroc boars between the ages of 590 and 630 days. Multiple linear regression analysis was performed to predict total and progressive motility and the total number of spermatozoa based on serum total L-carnitine and free L-carnitine levels. Total number of spermatozoa was not associated with basal serum L-carnitine levels. A regression equation was found in which both total L-carnitine levels and free L-carnitine levels were significant predictors of total and progressive motility (P 0.05). These results suggest that serum L-carnitine level is an important selection parameter for stock boars.

Source and level of dietary iron influence semen quality by affecting inflammation, oxidative stress and iron utilization levels in boars

Background Boars fed a mixed form of inorganic and organic iron in excess of the NRC recommended levels still develop anemia,which suggested that the current level and form of iron supplementation in boar diets may be inappropriate.Therefore,56 healthy Topeka E line boars aged 15–21 months were randomly divided into 5 groups:basal diet supplemented with 96 mg/kg ferrous sulfate(FeSO_(4))and 54 mg/kg glycine chelated iron(Gly-Fe,control);80 mg/kg or 115 mg/kg Gly-Fe;80 mg/kg or 115 mg/kg methionine hydroxyl analogue chelated iron(MHA-Fe,from CalimetFe)for 16 weeks.The effects of dietary iron supplementation with different sources and levels on semen quality in boars were investigated.Results 1)Serum Fe and hemoglobin concentrations were not affected by reduced dietary iron levels in the 80 mg/kg or 115 mg/kg Gly-Fe and MHA-Fe groups compared with the control group(P>0.05).2)Serum interleukin-6(IL-6)and sperm malondialdehyde(MDA)levels in the 80 mg/kg or 115 mg/kg MHA-Fe groups were lower than those in the control group(P<0.05),and higher serum superoxide dismutase levels and lower MDA levels in the 115 mg/kg MHA-Fe group(P<0.05).3)Boars in the 80 mg/kg or 115 mg/kg Gly-Fe and MHA-Fe groups had lower serum hepcidin(P<0.01),ferritin(P<0.05),and transferrin receptor(P<0.01)concentrations,and boars in the 115 mg/kg MHA-Fe group had higher seminal plasma Fe concentrations compared with the control group.4)Boars in the 80 mg/kg and 115 mg/kg MHA-Fe groups had lower abnormal sperm rate and in situ oscillating sperm ratio compared to the control group at weeks 12 and/or 16 of the trial.However,the effect of Gly-Fe on improving semen quality in boars was not evident.5)Serum IL-6 level was positively correlated with hepcidin concentration(P<0.05),which in turn was significantly positively correlated with abnormal sperm rate(P<0.05).Furthermore,significant correlations were also found between indicators of iron status and oxidative stress and semen quality parameters.Conclusions Dietary supplementation with 80 mg/kg or 115 mg/kg MHA-Fe did not induce iron deficiency,but rather reduced serum inflammatory levels and hepcidin concentration,alleviated oxidative stress,increased body iron utilization,and improved semen quality in adult boars.

Epizootiological surveillance of porcine circoviruses in free-ranging wild boars in China

Four species of porcine circoviruses(PCV1–4)have been reported to circulate in Chinese domestic pigs,while the epizootiology of these viruses in free-ranging wild boars in China remains unknown.In this study,tissue and serum samples collected from diseased or apparently healthy wild boars between 2018 and 2020 in 19 regions of China were tested for the prevalence of PCV1–4 infections.Positive rates of PCV1,PCV2,and PCV3 DNA in the tissue samples of Chinese wild boars were 1.6%(4/247),58.3%(144/247),and 10.9%(27/247)respectively,with none positive for PCV4.Sequence analysis of viral genome showed that the four PCV1 strains distributed in Hunan and Inner Mongolia shared 97.5%–99.6%sequence identity with global distributed reference strains.Comparison of the ORF2 gene sequences showed that 80 PCV2 strains widely distributed in 18 regions shared 79.5%–100%sequence identity with reference strains from domestic pigs and wild boars,and were grouped into PCV2a(7),PCV2b(31)and PCV2d(42).For PCV3,17 sequenced strains shared 97.2%–100%nucleotide identity at the genomic level and could be divided into PCV3a(3),PCV3b(2)and PCV3c(12)based on the phylogeny of ORF2 gene sequences.Serological data revealed antibody positive rates against PCV1 and PCV2 of 11.4%(19/167)and 53.9%(90/167)respectively.The data obtained in this study improved our understanding about the epidemiological situations of PCVs infection in free-ranging wild boars in China and will be valuable for the prevention and control of diseases caused by PCVs infection.

Preliminary attempt to distinguish the domesticated pigs from wild boars by the methods of carbon and nitrogen stable isotope analysis

Despite great achievements in the origins of domestic pigs made by the methods of zooarchaeology and molecular biology,how to scientifically distinguish the domesticated pigs from wild boars during the early stage of pig domestication is still poorly understood.Compared to wild boar's diets which come from the natural environment,the diets of domestic pigs are more easily influenced by human feeding activities.Therefore,in principle,exploration of the dietary differences among pigs and understanding the impact on pig diets fed by humans can have great potential to differentiate between wild boars and domesticated pigs.To reveal dietary differences among pigs and distinguish the domesticated pigs from wild boars based on comparison with the diets of humans and other animals,we analyzed the carbon and nitrogen stable isotopes of human bones from Xiaojingshan Site and animal bones from Yuezhuang Site,both of which belong to Houli Culture in Shandong Province and date to about 8500―7500 years ago.The mean δ 13C value((-17.8 ± 0.3)‰) and δ 15N value((9.0±0.6)‰) in human collagen indicate that although millet agriculture began it was not the main subsistence strategy as millets are typical of C4 plants and that humans made a living mainly by gathering,hunting or raising some domesticated animals.The δ 13C value(-16.1‰) and δ 15N value(6.9‰) in the bovine suggest that C3 plants were dominant in its diet with some C4 plants complemented.The fish has lower δ 13C value(-24.9‰) and higher δ 15N value(8.8‰) than the bovine,which is the characteristic of the isotopic values from Eurasian freshwater fish.Based on the differences in carbon and nitrogen isotope values,the pigs can be divided into three groups.A group,composed of two pigs,has low δ 13C values(-18.1‰,-20.0‰) and low δ 15N values(4.7‰,6.0‰).B group,only one pig,has the highest δ 13C value(-10.6‰) and mediate δ 15N value(6.4‰).As for the C group,also only one pig,low δ 13C value(-19.0‰) and the highest δ 15N value(9.1‰) are observed.Previous studies on the stable isotopes from modern or ancient wild boars' bones have suggested that C3 plants are predominated in their diets and that their δ 15N values are close to those in herbivores and far from those in carnivores.Based on the comparison with the isotope values from humans,the wild boars and the domestic pigs from Xipo Site in Henan 6000―5500 years ago and Kangjia Site in Shaanxi 4500―4000 years ago,we conclude that A pig group belongs to wild boars while B and C groups can be attributed to domesticated pigs.

Glutamate and aspartate alleviate testicular/epididymal oxidative stress by supporting antioxidant enzymes and immune defense systems in boars

Several potential oxidative agents have damaging effects on mammalian reproductive systems.This study was conducted to investigate the effects of glutamate(Glu)and aspartate(Asp)supplementation on antioxidant enzymes and immune defense systems in the outer scrotum of boars injected with H2O2.A total of 24 healthy boars were randomly divided into 4 treatment groups:control(basal diet,saline-treated),H2O2(basal diet,H2O2-challenged outer scrotum(1 m L kg^–1 BW)),Glu(basal diet+2%Glu,H2O2-challenged),and Asp(basal diet+2%Asp,H2O2-challenged).Our results showed that both Glu and Asp supplementation improved testicular morphology and decreased the genital index in the H2O2-treated boars.Glu and Asp administration increased the antioxidant enzyme activities and affected the testicular inflammatory cytokine secretion but had no effect on sex hormone levels.Furthermore,the m RNA expression of CAT,Cu Zn SOD,and GPx4 was altered in the testes and epididymis of boars treated with Asp and Glu.Glu and Asp supplementation also modulated the expression of TGF-β1,IL-10,TNF-α,IL-6 and IL-1βin the testis and epididymis.These results indicate that dietary Glu and Asp supplementation might enhance antioxidant capacity and regulate the secretion and expression of inflammatory cytokines to protect the testes and epididymis of boars against oxidative stress.

Pig Reproductive and Respiratory Syndrome Virus (PRRSV) Does Not Attach to Boar Sperm;It Affects Only the Velocity Pattern and the Mobility Pattern

The purpose of the study was to evaluate the sperm viability of semen infected with PRRSV viral particles, observing the effect of the Virus on the motility of boar sperm. The work was carried out at the FMVZ-BUAP Genetics and Reproduction Laboratory. 5 stallions were used. Each sample contained 1 × 106 sperm, the PRRS virus strain was ATCC-VR-2332 (0, 102, 104 and 106 copies of RNA/mL in triplicate), it was observed daily at the CASA;Hamilton Thorne®. Cells with MT (P < 0.05) on days 1, 3, 5, 7 and 10 of evaluation with 201 ± 7.3, 167 ± 10.1, 165 ± 14.6, 134 ± 8.2 and 120 ± 8.8, respectively. The % MP between control and virus concentrations (P ≥ 0.05). The LCV on day 1 and 7 PI at 10X2 and 10X6 (P < 0.05) vs control. In the Correlation Matrix, where it is observed that there is a correlation between VSL and VAP, VSL and VCL, VCL and ALH, VAP with ALH. There is a correlation of VSL and ALH, STR and ALH. In this study there were (P ≤ 0.01) in the VCL, in the concentrations (102) 162.81 ± 10.65 and (106) 177.12 ± 5.77 vs 193.04 ± 4.62 of control. This indicates that altering these parameters would be related to fertility and the PRRS virus affects the LCV. Regarding the VSL, it was observed that the sperm infected with viruses 102, 104 and 106 of 48.00 ± 3.38, 49.88 ± 1.83 and 50.55 ± 2.24 Vs. 56.66 ± 1.68 of control respectively, the control would have greater possibilities of fertilizing the oocyte. In this study, it was found (P ≤ 0.01) in the VAP with 102 of 77.26 ± 5.16, 104 with 83.35 ± 2.41 and 106 with 81.29 ± 3.14 vs the control with 90.56 ± 2.07. Regarding the ALH there is (P < 0.05) a 104 with 8.70 ± .26 and 106 with 9.64 ± 0.23 vs control 8.50 ± 0.27. The presence of different concentrations of PRRSV in boar semen induces changes in different types of sperm motility. Infection of ejaculates with the PRRS virus affects sperm motility on days 1, 3, 5, 7, and 10 post-infections.

Investigation on the Harm of Wild Boar in Mingtong Town, Chengkou, Chongqing

Through the investigation and analysis of the wild boars in Mingtong Town, Chengkou, Chongqing, it was concluded that the area had long been seri- ously affected by wild boars and some of the human activities were potentially threatened, and the potential threat areas were identified. Corresponding suggestions were proposed to guide the prevention and control work, so as to protect the habi- tat and reproduction of wildlife and ensure the normal production and life of the masses.

Effects of DHA-enriched hen egg yolk and L-cysteine supplementation on quality of cryopreserved boar semen

The objective of the present study was to determine the effects of docosahexaenoic acid （DHA）-enriched hen egg yolks and L-cysteine supplementation on the qualities of the cryopreserved boar semen. A total of 15 ejaculates from 5 Pietrain boars were divided into 4 groups according to the compositions of the freezing extenders used, that is, normal hen egg yolk （group Ⅰ）, DHA-enriched hen egg yolk （group Ⅱ）, normal hen egg yolk with 5 mmol L^-1 of cysteine supplementation （group Ⅲ） and DHA-enriched hen egg yolk with 5 mmol L1 of cysteine supplementation （group Ⅳ）. The semen was cryopreserved using controlled rate freezer and was thawed at 50℃ for 12 s. Progressive motility, sperm viability, acrosome integrity and functional integrity of sperm plasma membrane of the post-thawed semen were evaluated. The supplementation of L-cysteine in the freezing extender alone （group Ⅲ） improved progressive motility （P 〈 0.05）, and the supplementation of L-cysteine in combination with DHA-enriched hen egg yolk （group Ⅳ） improved both progressive motility （P 〈 0.05） and acrosome integrity （P 〈 0.01）. The use of DHA-enriched hen egg yolk alone （group Ⅱ） did not enhance any of the post-thawed semen qualities （P 〉 0.05）. In conclusion, the supplementation of antioxidant L-cysteine alone or in combination with DHA-enriched hen egg yolk significantly improved the post-thawed semen qualities, especially progressive motility and acrosome integrity.

Supplemental effect of varying L-cysteine concentrations on the quality of cryopreserved boar semen

Cryopreservation is associated with the production of reactive oxygen species, which leads to lipid peroxidation of the sperm membrane and consequently a reduction in sperm motility and decreased fertility potential. The aim of this study was to determine the optimal concentration of L-cysteine needed for cryopreservation of boar semen. Twelve boars provided semen of proven motility and morphology for this study. The semen was divided into four portions in which the lactose-egg yolk （LEY） extender used to resuspend the centrifuged sperm pellet was supplemented with various concentrations of L-cysteine to reach 0 mmol L^-1 （group Ⅰ, control）, 5 mmol L^-1 （group Ⅱ）, 10 mmol L^-1 （group Ⅲ） and 15 mmol L^-1 （group Ⅳ）. Semen suspensions were loaded in straws （0.5 mL） and placed in a controlled-rate freezer. After cryopreservation, frozen semen samples were thawed and investigated for progressive motility, viability using SYBR-14/EthD-1 staining and acrosome integrity using FITC-PNA/EthD-1 staining. There was a significantly higher （P 〈 0.01） percentage of progressive motility, viability and acrosomal integrity in two L-cysteine-supplemented groups （group Ⅱ and group Ⅲ） compared with the control. There was a biphasic effect of L-cysteine, with the highest percentage of progressive motility, viability and acrosomal integrity in group Ⅲ. In conclusion, 5 or 10 mmol L^-1 was the optimum concentration of L-cysteine to be added to the LEY extender for improving the quality of frozen-thawed boar semen.

GSTM3, but not IZUMO1, is a cryotolerance marker of boar sperm

Background: Cryopreservation is currently the most efficient method for long-term preservation of mammalian gametes and is extensively used in swine artificial insemination(AI) centres. However, it is well-known that cryopreservation procedures induce changes in the water phase in both intra and extracellular compartments,which alter the content and localisation of several proteins and ends up curtailing the structural integrity of functional sperm(i.e., cryoinjuries). Alterations and deficiencies of sperm-oocyte binding proteins during gamete recognition are one of the causes of reproductive failure both in vitro and in vivo. In this sense, characterisation of cryopreservation effects upon oocyte-binding proteins of sperm, such as IZUMO1 and GSTM3, is essential when assessing the impact of this technique in swine reproduction.Results: Cryopreservation was found to induce changes in the localisation of IZUMO1 and GSTM3 in boar sperm.However, the relative content of both proteins was not altered after thawing. Furthermore, whereas IZUMO1 content was found not to be related to the cryotolerance of boar sperm, GSTM3 content was observed to be higher in poor(PFE) than in good(GFE) freezability ejaculates in both pre-frozen(1.00 INT·mm^2± 0.14 INT·mm^2 vs.0.72 INT·mm^2± 0.15 INT·mm^2;P < 0.05) and post-thawed(0.96 INT·mm^2± 0.20 INT·mm^2 vs. 70 INT·mm^2± 0.19 INT·mm^2;P < 0.05) samples. Moreover, GSTM3 levels were found to be higher in those spermatozoa that exhibited low mitochondrial activity, high reactive oxygen species(ROS) production, and high membrane lipid disorder postthaw(P < 0.05).Conclusions: The difference in GSTM3 content between GFE and PFE, together with this protein having been found to be related to poor sperm quality post-thaw, suggests that it could be used as a cryotolerance marker of boar spermatozoa. Furthermore, both IZUMO1 and GSTM3 relocate during cryopreservation, which could contribute to the reduced fertilising capacity of frozen-thawed boar sperm.