Controlling contagious agalactia in artificial insemination centers for goats and detection of Mycoplasma mycoides subspecies capri in semen

Many goat artificial insemination (AI) centers in Spain have adopted new measures to control contagious agalactia (CA). To avoid the introduction of male goats carrying mycoplasma organisms subclinically in their external ear canal (auricular carriers) in these centers, two ear swabs and a blood sample are obtained from all candidate animals for polymerase chain reaction (PCR), culture (swabs) and serologic tests to detect the presence of mycoplasmas. In addition, the semen produced at these centers is routinely cultured and PCR tested also to detect the presence of mycoplasmas. One y after the introduction of this program, we tested 48 ear swabs and 24 blood samples from 24 candidates for admission to these AI Centers. Three of these ear swab samples (3/48, 6.25%) scored positive for the presence of mycoplasmas; Mycoplasma agalactiae (Ma) was detected in two samples and Mycoplasma mycoides subsp. capri (Mmc) in one. All animals were serologically negative for Ma. Also, out of 173 semen samples obtained from 137 admitted animals (2 and 3 samples were obtained in 16 and 10 bucks, respectively), one (1/173, 0.56%) was positive for Mmc. Our findings suggest that ear swab and semen samples are useful tools to control CA at AI Centers. The introduction of this program has also resulted in the first detection of Mmc in semen from a naturally infected goat, confirming the ability of this mycoplasma to colonize the reproductive tract of male goats. These results highlight the need to improve control measures in semen producing centers to minimize the risk of CA transmission.     

Presence of contagious agalactia causing mycoplasmas in Spanish goat artificial insemination centres

Male goats admitted to artificial insemination centres come from herds that have shown no clinical symptoms of contagious agalactia (CA) for the last 6 mo. However, prior reports suggest that this control measure may not be completely effective. This study was designed to detect the presence of CA-causing mycoplasmas in 9 Spanish centres, comprising 159 goats (147 males and 12 teaser does) of 8 different breeds. A microbiological study was conducted during 8 mo on 448 samples (318 ear swabs, 119 semen samples and 11 milk samples). In 86 samples (84 swabs, 1 semen sample and 1 milk sample), CA-causative mycoplasmas were detected by PCR or culture, and 52 animals (49 goat males and 3 teaser does) tested positive. Most of these positive animals were auricular carriers (n = 50), mainly of Mycoplasma mycoides subsp. capri (Mmc), although some M. agalactiae (Ma) and, interestingly, M. capricolum subsp. capricolum (Mcc) carriers were also identified. At least 1 animal infected by CA-causing mycoplasmas was detected in 8 of the 9 centres (88.8%) although in most (66.7%) no infected animals or only 1 or 2 positive animals were identified. Our results indicate the presence of CA carriers as asymptomatic animals in reproductive programmes. These findings have already prompted efficient measures to detect and avoid the entry of these carriers in Spanish centres. We recommend similar measures for all centres in areas where CA is endemic.     

Surveillance of Mycoplasma agalactiae and Mycoplasma mycoides subsp. capri in dairy goat herds

This study was designed to monitor the presence of Mycoplasma agalactiae and Mycoplasma mycoides subsp. capri (Mmc) in 66 dairy goat herds of a genetic improvement programme in a region of Spain where contagious agalactia is endemic. Over a whole lactation period, 300 bulk tank milk and 381 milk samples from goats with clinical mastitis were subjected to polymerase chain reaction (PCR) to detect the two mycoplasma species. The presence of mycoplasmas (either species or both) was detected in 66.7% of the herds and M. agalactiae was identified in 95.45% of these positives herds. In a given infected herd, mycoplasmas were not continuously detected over the whole study period. Our findings indicate that in an endemic area, M. agalactiae and Mmc can be monitored through PCR analysis of mastitic milk and bulk tank milk (BTM) samples. Over a lactation period we recommend testing multiple BTM samples on a herd. No relationship was observed between the use of inactivated mycoplasma vaccines and the PCR detection of both mycoplasmas.     

Detection of Leptospira spp. in semen and vaginal fluids of goats and sheep by polymerase chain reaction

Thirteen goat herds and seven sheep flocks in the state of Rio de Janeiro, Brazil were screened for leptospirosis. From the three herds and three flocks with greatest seroreactivity, 19 goats (16 females and three bucks) and 40 sheep (26 ewes and 14 rams) that were seropositive (specific anti-Leptospira titres > or =400, based on a microscopic agglutination test), were selected for more detailed studies. From those animals, samples of vaginal fluids or semen were collected for bacteriological and molecular assays. For both species of animals, the most prevalent reactions were to serovars Hardjo, Shermani, and Grippotyphosa. Although leptospires were detected by darkfield microscopy in three vaginal fluid samples (from two goats and one ewe), pure isolates were not obtained by bacteriological culture of vaginal fluids or semen. However, seven vaginal fluid samples (from four goats and three ewes) and six semen samples (all from rams) were positive on polymerase chain reaction (PCR). Based on these findings, in addition to analogous findings in cattle, we inferred that there is potential for venereal transmission of leptospirosis in small ruminants.     

Effects of lactobacillus plantarum ZJ316 on pig growth and pork quality

Background

Contagious agalactia (CA) of sheep and goats caused by Mycoplasma agalactiae is a widely occurring economically important disease that is difficult to control. The ELISA is commonly used for the serological detection of CA but it has some limitations and the performance of the available tests have not been properly evaluated. Two commercial ELISA kits are widely used, one involving a fusion protein as target antigen and the other a total antigen. The objectives were to compare these tests by evaluating: i. Their diagnostic sensitivity and specificity, the relevance of the recommended cut-off points, the correlation between the two tests, and, the correlation between serology data and the milk shedding of M. agalatiae; ii. The influence of extrinsic factors such as the targeted animal species, geographical origin of the samples, intra-specific variability of M. agalactiae and concurrent mycoplasma infections. A sample of 5900 animals from 211 farms with continuous CA monitoring for 20?years and no prior vaccination history was used. The infection status was known from prior bacteriological, epidemiological and serological monitoring with a complementary immunoblotting test.

Results

The average diagnostic sensitivity was 56% [51.8?C59.8] for the fusion protein ELISA and 84% [81.3?C87.2] for the total antigen ELISA, with noteworthy flock-related variations. The average diagnostic specificity for the fusion protein ELISA was 100% [99.9?C100], and for the total antigen ELISA differed significantly between goats and sheep: 99.3% [97.4?C99.9] and 95.7% [93.8?C97.2] respectively. Experimental inoculations with different M. agalactiae strains revealed that the ELISA kits poorly detected the antibody response to certain strains. Furthermore, test performances varied according to the host species or geographical origin of the samples. Finally, the correlation between milk shedding of M. agalactiae and the presence of detectable antibodies in the blood was poor.

Conclusions

These serological tests are not interchangeable. The choice of a test will depend on the objectives (early detection of infection or disease control program), on the prevalence of infection and the control protocol used. Given the variety of factors that may influence performance, a preliminary assessment of the test in a given situation is recommended prior to widespread use.     

Flaxseed oil modulates testicular biometrics,hormone, libido,antioxidant and semen profiles in endangered Teressa goat of Andaman and Nicobar Islands

Teressa goat is a unique goat breed in Andaman and Nicobar Islands (ANI) of India. Effects of Flaxseed oil (FSO) supplementation in body weight (BW), scrotal circumference (SC), testicular volume (TV) and testicular weight (TW), endocrinological profiles, sex behavioural profiles (SBPs), oxidative stress markers and semen production and its quality profiles in rainy and dry summer season were studied in Teressa goat. Male goats (n = 12) of 3–4 years old were equally divided into control and treated groups. Treated animals received 25 mL FSO per day. Oral drenching of FSO was done in the morning before feeding the concentrate ration. Body weight, scrotal circumference, TV and TW were measured in bucks of FSO treated and untreated during rainy and dry summer seasons. Blood follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone, thyroid stimulating hormone (TSH), triiodothyronine (T3), thyroxine (T4), cortisol and prolactin, total antioxidant capacity (TAC), superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and malondialdehyde (MDA) were measured in bucks of FSO treated and untreated during rainy and dry summer seasons. Libido score (LS), mating ability score (MAS) and sex behavioural score (SBS) were estimated at time of semen collection in bucks of FSO treated and untreated during rainy and dry summer seasons. Semen samples (n = 100; 50 semen samples from each season; each 25 semen samples from control and treatment groups per season) were collected and analysed for semen quality profiles. One-way ANOVA (control rainy, control dry, treated rainy and treated dry) revealed that BW, SC, TV and TW, FSH, LH, testosterone, TSH, T3 and T4 were higher (P < 0.05) and cortisol and prolactin were lower (P < 0.05) in FSO treated bucks of rainy season followed by untreated bucks of rainy season, FSO treated bucks of dry summer season and were lower (P < 0.05) in untreated bucks of dry summer season. Similarly, TAC, CAT, SOD and GSH, LS, MAS and SBS, and volume, pH, sperm concentration, mass activity, total motility (TM), viability, acrosomal integrity (AcI), plasma membrane integrity (PMI) and nuclear integrity (NI) were higher (P < 0.05) and MDA and TSA were lower (P < 0.05) in FSO treated bucks of rainy season followed by FSO treated bucks of dry summer season, untreated bucks of rainy season and were lower (P < 0.05) in untreated bucks of dry summer season. The results of the present study indicated that the breeding bucks suffered physiological stress (higher cortisol), oxidative stress (higher MDA and deficiency of antioxidants), hormonal imbalance (higher prolactin and cortisol and deficiency of gonadotropins, gonadal hormone and thyroid hormones) and infertility due to poor libido and poor semen production and its quality profiles during dry summer season. Thus, dry summer was more stressful season compared to rainy season for the goat bucks. FSO supplementation mitigated these stresses and improved the scrotal and testicular biometrics, libido, antioxidants, hormones and semen quality profiles in Teressa goat bucks. The current study concluded that FSO effectively improved the hormones, libido, antioxidant profiles, and scrotal and testicular biometrics with cascading beneficial effects on semen quality profiles in Teressa goat bucks under humid tropical island ecosystem of Andaman and Nicobar Islands.     

The effects of cryopreservation on the morphometric dimensions of caprine sperm heads

Cryopreserved semen has been utilised in the artificial insemination of livestock species for over 40 years, even though the detrimental effects of cryopreservation on sperm function and fertility are well documented. In the present study, computer-automated sperm-head morphometry was used to determine if goat sperm-head morphometry was affected by freezing and thawing. A microscope slide was prepared from single semen samples, collected by artificial vagina, from 10 sexually active Saanen bucks. The remainder of each sample was frozen in a tris-citrate-yolk extender. After thawing, semen smears were prepared on microscope slides. All slides were stained in haematoxylin and mean sperm-head measurements of length, width, width/length, area and perimeter were determined for each slide by computer aided sperm morphometry analysis. The effects of sperm freezing on sperm-head dimensions within and among all bucks were determined. No significant (P > 0.10) freezing effect was found between fresh semen and postthaw samples for length (7.00 μm vs 7.13 μm), width (3.77 μm vs 3.87 μm), width/length (0.54 μm vs 0.54 μm), area (19.67 μm² vs 20.57 μm²) and perimeter (18.62 μm vs 18.83 μm) when analysed across all bucks. Significant differences (P < 0.05) were however found within three bucks for area, perimeter, length and width, with the percentage increase in measurements being significantly greater than in the remaining bucks. The variability of the morphometric dimensions were not affected by freezing. The results indicate that semen freezing did not affect the overall dimensions of sperm heads across the entire population of bucks sampled. However, since sperm-head dimensions from three bucks were affected, changes in sperm-head morphometry may be indicative of spermatozoa of the semen from individuals to successfully freeze. Because the overall mean sperm-head dimensions acquired from frozen/thawed semen were not different from those of fresh semen, previously reported measurements of goat sperm heads are probably reflective of fresh semen. More importantly, retrospective studies of sperm-head morphometry and fertility may now be performed utilising extensive breeding records from frozen semen.     

Profile of the Caprine arthritis-encephalitis virus (CAEV) in blood, semen from bucks naturally and experimentally infected in the semi-arid region of Brazil

The aim of this study was to report the chronology of Caprine arthritis-encephalitis virus elimination and compare the blood and semen viral profiles of animals naturally and experimentally infected by SRLV raised in the semi-arid region of Brazil. The experiment was carried out at the Brazilian Center for Goat Research (Embrapa). Nine bucks were selected, four naturally infected by CAEV and five animals proven negative that were inoculated with the goat lentivirus (CAEV-Cork strain). Every week the animals were submitted to semen collection using an artificial vagina. The blood was collected by puncturing the jugular vein with tubes containing EDTA, 7 days after inoculation (experimentally infected group) or at the start of the experiment (naturally infected group) and then at every 30 days. The genomic viral DNA was extracted from semen and blood and then Nested-PCR was applied. An agar gel microimmunodiffusion was performed to detect anti-CAEV antibodies. The results were described in percentage and analyzed by the Chi square test (P < 0.05). The presence of anti-CAEV antibodies was detected in the 16th week after inoculation that characterized the seroconversion from four of the five naturally infected goat bucks (80%). The fifth reproducer presented late seroconversion, totaling 32 weeks post-inoculation. A quantity was observed in the total of samples collected of 12.50 and 17.14% positive results in the blood and 10.98 and 11.25% positive results in the semen of the naturally and experimentally infected animals, respectively, and there was no statistical difference. No statistically significant differences were observed regarding the presence of proviral DNA in the blood and semen of the naturally and experimentally infected animals. A viral elimination pattern was not identified during the assessment period, but the presence of proviral DNA was shown at shorter intervals after the 18th week and the 22nd week, for the experimentally and naturally infected bucks, respectively. Therefore, recently infected goats in the period prior to seroconversion eliminated small ruminant lentivirus proviral DNA in the semen and are important sources of infection that should be considered in a control program of this lentivirus, and the Nested-PCR technique is a relevant tool to select virus-free ejaculates.     

Is caprine arthritis encephalitis virus (CAEV) transmitted vertically to early embryo development stages (morulae or blastocyst) via in vitro infected frozen semen?

The aim of this study was to determine, in vivo, whether in vitro infected cryopreserved caprine sperm is capable of transmitting caprine arthritis-encephalitis virus (CAEV) vertically to early embryo development stages via artificial insemination with in vitro infected semen. Sperm was collected from CAEV-free bucks by electroejaculation. Half of each ejaculate was inoculated with CAEV-pBSCA at a viral concentration of 10⁴ TCID₅₀/mL. The second half of each ejaculate was used as a negative control. The semen was then frozen. On Day 13 of superovulation treatment, 14 CAEV-free does were inseminated directly into the uterus under endoscopic control with thawed infected semen. Six CAEV-free does, used as a negative control, were inseminated intrauterine with thawed CAEV-free sperm, and eight CAEV-free does were mated with naturally infected bucks. Polymerase chain reaction (PCR) was used to detect CAEV proviral-DNA in the embryos at the D7 stage, in the embryo washing media, and in the uterine secretions of recipient does. At Day 7, all the harvested embryos were PCR-negative for CAEV proviral-DNA; however, CAEV proviral-DNA was detected in 8/14 uterine smears, and 9/14 flushing media taken from does inseminated with infected sperm, and in 1/8 uterine swabs taken from the does mated with infected bucks. The results of this study confirm that (i) artificial insemination with infected semen or mating with infected bucks may result in the transmission of CAEV to the does genital tack seven days after insemination, and (ii) irrespective of the medical status of the semen or the recipient doe, it is possible to obtain CAEV-free early embryos usable for embryo transfer.     

Sedation or anaesthesia decrease the stress response to electroejaculation and improve the quality of the collected semen in goat bucks

Electroejaculation (EE) is stressful and probably painful; thus the administration of anaesthesia is recommended to decrease those negative effects. However, anaesthesia has a direct risk of provoking death, but sedation is less risky than anaesthesia. At the same time, α2-adrenergic agonists may improve semen quality. Therefore, the aim of the present study was to compare the physiological and behavioural responses indicative of stress and possibly pain, and the semen quality in electroejaculated untreated, anaesthetised or sedated goat bucks. Semen was collected from eight bucks using three different procedures in all them (EE in untreated bucks, EE under sedation or EE under general anaesthesia). The number of vocalizations during EE and the behavioural pattern before and after procedures were recorded. Pain visual analogue scale (VAS) score was also determined during EE. Rectal temperature, heart rate, serum cortisol concentration, biochemical and haematological parameters were measured before and after each procedure, and sperm characteristics were determined. Bucks vocalised more often when untreated than sedated or anaesthetised (P<0.02). The pain VAS score was greater when bucks were untreated than sedated or anaesthetised (P<0.002). The rectal temperature, heart rate, total protein, albumin and haemoglobin concentrations were greater when bucks were untreated than anaesthetised or sedated (P<0.02). Serum cortisol increased after EE (P=0.0006), without differences between procedures. The frequency and duration of lying down after EE were greater when bucks were anaesthetised than sedated or untreated (P<0.05), and were also greater when bucks were sedated than untreated (P<0.05). The number of times that the animal tried to stand up after EE was greater when bucks were anaesthetised than sedated or untreated (P<0.02). The sperm mass motility was greater when bucks were anaesthetised or sedated than when they were untreated (P=0.048). When animals were sedated, the ejaculate contained more spermatozoa with functional plasma membrane (P=0.03) and morphologically normal (P=0.05) than when they were untreated. In conclusion, general anaesthesia and sedation decreased the stress and probably the pain response provoked by EE and especially sedation improved the quality of the semen collected.     

Undernutrition reduces the body weight and testicular size of bucks exposed to long days but not their ability to stimulate reproduction of seasonally anestrous goats

In semiarid conditions, feed is often scarce and variable with underfeeding being common; these factors can potentially induce fertility reductions in both sexes. Sexually active bucks are able to very efficiently fertilize out-of-season goats, but we do not know whether underfeeding would reduce the ability of bucks to fertilize goats during these periods. Two experiments were conducted to determine (i) testicular size and change of odor intensity of undernourished bucks exposed to long days and (ii) the ability of these bucks to stimulate reproductive activity in seasonally anestrous goats. In experiment 1, bucks (n = 7) were fed 1.5 times the normal maintenance requirements from September to May and formed the well-fed group. Another group of bucks (n = 7) were fed 0.5 times the maintenance requirements and formed the undernourished group. All bucks were subjected to artificially long days from 1 November to 15 January; this period was followed by a natural photoperiod until 30 May. Body weight, scrotal circumference and male odor intensity changes were determined every 2 weeks. In experiment 2, two groups of female goats (n = 26 each) were exposed to well-fed (n = 2) or undernourished bucks (n = 2) on 31 March. Ovulations and pregnancy rates were determined by transrectal ultrasonography. In experiment 1, a treatment by time interaction was detected for BW, scrotal circumference and odor intensity changes (P < 0.001). The BWs of well-fed bucks were greater than those of the undernourished bucks from October to May (P < 0.01), as were the scrotal circumferences from December to March (P < 0.05) and odor intensities from February to May (P < 0.05). In experiment 2, the proportions of females that ovulated at least once (100% v. 96%) or those that were diagnosed as pregnant (85% v. 77%; P > 0.05) did not differ significantly between the goats exposed to well-fed or undernourished bucks. The interval between the introduction of bucks and the onset of estrous behavior was shorter in goats exposed to well-fed bucks compared to the interval for those goats exposed to undernourished bucks (2.5 ± 0.2 v. 9.5 ± 0.6 days; P < 0.05). We conclude that undernourishment reduces the testicular size and odor intensity responses in bucks exposed to long days, but that undernourished bucks are still able to stimulate reproductive activity in seasonally anestrous goats, as is also the case for well-fed bucks.     

Seasonal changes in sperm DNA fragmentation of Murciano-Granadina goats: The compelling case for dynamic assessment

Evidence is provided of seasonal changes in the rate of DNA fragmentation dynamics of sperm collected from goats Murciano-Granadina breed), in a Spanish Breeding Centre at 41°N latitude. Results show that the initial assessment of sperm DNA fragmentation (T0) was not sufficient to differentiate sperm DNA fragmentation of bucks collected at different times of the year (Kruskal–Wallis 2.399; P = 0.493). However, when thawed semen was subsequently incubated at 37 °C for periods of between 2 and 48 h, differences were recorded in the rate of sperm DNA fragmentation (r-SDF), that are indicative of seasonal changes in reproduction (Log Rank: Mantel–Cox; χ² = 158.6; DF = 3; P < 0.001). Goat semen collected and cryopreserved in the Spanish winter and spring resulted in a poor post-thaw sperm DNA longevity and recorded a r-SDF of 0.8 and 1.3 per hour, after 48 h of incubation respectively. The r-SDF subsequently decreased to 0.16 and 0.36 per hour during summer and autumn. It is therefore, recommended that semen samples from Murciano-Granadina goats housed above the 40°N latitude be collected and cryopreserved in the summer and early autumn. This would result in a more stable DNA molecule, which is likely to improve the reproductive success.     

Mycoplasma agalactiae MAG_5040 is a Mg2+-Dependent,Sugar-Nonspecific SNase Recognised by the Host Humoral Response during Natural Infection

In this study the enzymatic activity of Mycoplasma agalactiae MAG_5040, a magnesium-dependent nuclease homologue to the staphylococcal SNase was characterized and its antigenicity during natural infections was established. A UGA corrected version of MAG_5040, lacking the region encoding the signal peptide, was expressed in Escherichia coli as a GST fusion protein. Recombinant GST-MAG_5040 exhibits nuclease activity similar to typical sugar-nonspecific endo- and exonucleases, with DNA as the preferred substrate and optimal activity in the presence of 20 mM MgCl₂ at temperatures ranging from 37 to 45°C. According to in silico analyses, the position of the gene encoding MAG_5040 is consistently located upstream an ABC transporter, in most sequenced mycoplasmas belonging to the Mycoplasma hominis group. In M. agalactiae, MAG_5040 is transcribed in a polycistronic RNA together with the ABC transporter components and with MAG_5030, which is predicted to be a sugar solute binding protein by 3D modeling and homology search. In a natural model of sheep and goats infection, anti-MAG_5040 antibodies were detected up to 9 months post infection. Taking into account its enzymatic activity, MAG_5040 could play a key role in Mycoplasma agalactiae survival into the host, contributing to host pathogenicity. The identification of MAG_5040 opens new perspectives for the development of suitable tools for the control of contagious agalactia in small ruminants.     

Comparison of semen collection techniques in goats

Characteristics of goat semen collected with the artificial vagina (AV), electroejaculator (EE), and Bailey ejaculator (BE) were compared. Semen was collected three times by each method from four bucks for a total of 36 separate collections. The semen was evaluated for volume, sperm concentration, mass activity (0 to 4, no activity to rapid wave motion), sperm motility (%), pH, and acrosome morphology (% normal). The means (± SEM) of semen samples collected with the AV, EE, and BE were volume-0.4 (±0.1), 0.7 (± 0.2), 0.6 (± 0.1) ml; concentration-4.6 (± 0.5), 2.1 (± 0.3), 1.4 (± 0.2) 10⁹/ml; mass activity-4.0 (± 0), 3.3 (± 0.3), 2.8 (± 0.2); motility-81 (± 2), 78 (± 2), 76 (± 2) %; pH-6.2 (± 0.1), 7.0 ± (0.2), 7.0 ± (0.2); and normal acrosomes-96 (± 1), 92 (± 3), and 88 (± 3) %, respectively. Semen collected with the AV was lower in volume than that collected with the BE (P < 0.05). The sperm concentration in semen samples collected with the AV was greater than those collected with the EE and BE (P < 0.05). Mass activity was greater and pH was less in AV samples than in EE or BE samples (P < 0.05). There was no difference in sperm motility of semen samples collected by the AV, EE, or BE. The average percentage of normal acrosomes was greater for AV than for BE samples (P < 0.05). Use of the EE and especially the BE resulted in increased vocalization by the goats and excessive muscular contractions of the rear limbs. The BE, in its present form (11.2 volts), is not recommended for semen collection in goats.     

Physiological response and semen quality of rabbit bucks supplemented with Moringa leaves ethanolic extract during summer season

Exposure of rabbit bucks to summer heat stress reduces their homeostasis and semen quality leading to a temporal subfertility. The potentiality of ethanolic extract of Moringa oleifera leaves (M. oleifera ethanolic extract (MLEE)) to reduce negative impacts of heat stress on physiological and semen quality traits was investigated. A total of 28 adult V-line rabbit bucks were randomly distributed among four experimental groups of seven rabbits each. The first group received water (placebo) and served as a control (M0). The other three groups were given orally MLEE at levels of 50 (M50), 100 (M100) and 150 (M150) mg/kg BW every other day for 12 consecutive weeks during the summer season. Chemical constituents of MLEE were detected by gas chromatography/MS. During the experimental period, ambient temperature and relative humidity were recorded daily and were used to estimate temperature and humidity index. Feed intake, BW, rectal temperature were recorded and blood serum biochemical attributes were determined. Semen samples were collected weekly and were analyzed for semen quality traits. Results showed that MLEE contained high percentages of long-chain fatty acids and antioxidant agents. Feed intake and BW were not affected significantly by the treatment, however rectal temperature was decreased significantly by 0.42°C, 0.24°C and 0.40°C in the M50, M100 and M150 groups, respectively, compared with the M0 group. Treatment with 50 mg/kg BW increased concentration of serum albumin (115%; P<0.05), total antioxidant capacity (132%; P<0.05) and testosterone (160%; P=0.098) as well as seminal plasma initial fructose (127%; P=0.092) compared with the control group. Compared with the control, MLEE supplementation with 50, 100 and 150 mg/kg BW increased significantly sperm concentration by 118%, 151% and 158%, sperm progressive motility by 117%, 120% and 118%, sperm viability by 129%, 137% and 127%, sperm normal morphology by 114%, 113% and 114%, intact acrosome sperm by 109% (on average) and sperm with integrated cell membrane by 109%, 123% and 114%, respectively. In conclusion, MLEE supplementation at a level of 50 mg/kg BW could be effectively used to improve heat tolerance, oxidative status and semen quality of rabbit bucks during summer season.     

Adverse effects of members of the Enterobacteriaceae family on boar sperm quality

Semen samples collected in 2012 from 1785 boars belonging to five different breeds were recruited from the quality control laboratory of Magapor SL, Spain. These samples came from 43 boar studs and resulted from diluting the ejaculates in commercial semen extenders. Evaluation of the semen sample characteristics (color, smell, pH, osmolality, concentration, motility of sperm cells, agglutination, acrosome integrity, short hypoosmotic swelling test, and abnormal forms) revealed that they met the international standards. The samples were also tested for the presence of aerobic bacterial contamination. In the present study, 14.73% (n = 263) of the semen samples were contaminated above 3 × 10² colony-forming units/mL with at least one type of bacteria. The Enterobacteriaceae family was by far the major contaminant, being present in 40.68% of the contaminated samples (n = 107). Bacterial strains of the Enterobacteriaceae family isolated from boar semen samples were in order of incidence (percentage of the contaminated samples): Serratia marcescens (12.55%), Klebsiella oxytoca (11.79%), Providencia stuartii (9.12%), Morganella morganii (3.80%), Proteus mirabilis (1.90%), and Escherichia coli (1.52%). We have seen that the presence in semen samples of S. marcescens, K. oxytoca, M. morganii, or P. mirabilis, but not P. stuartii or E. coli, was negatively associated with sperm motility (P < 0.05). The mean sperm concentration (P < 0.05), the mean percentage of spermatozoa with curled tails after the short hypoosmotic swelling test (P < 0.01), and the incidence of morphologically normal acrosomes (P < 0.05) were also lower in semen samples infected with M. morganii compared with uninfected ones. Moreover, P. mirabilis was negatively associated with the presence of abnormal forms. Thus, on the basis of the pathological effects that some of these strains may have on boar sperm quality, bacterial contamination should always be examined in semen samples prepared for artificial insemination.     

Caprine mycoplasmal mastitis in Nigeria

In a study to investigate the current status of intramammary mycoplasmosis in caprine udders in Nigeria, a total of 57 and 24 milk samples were collected from udders of goats affected by mastitis and from apparently normal goats’ udders, respectively. Acute and chronic mastitis were more commonly observed in goats between 1 and 3 years old. Mycoplasma agalactiae and Mycoplasma capricolum occurred at a significantly higher rate (p<0.05) in udders affected by mastitis than in normal healthy udders. Other mycoplasma occurring in low prevalence include Mycoplasma bovis and Mycoplasma mycoides subsp. mycoides LC. It is concluded that cultural (microbiological) surveillance is necessary for effective treatment and control of the disease in Nigeria.     

Real-Time PCR Investigation of Potential Vectors, Reservoirs, and Shedding Patterns of Feline Hemotropic Mycoplasmas

Three hemotropic mycoplasmas have been identified in pet cats: Mycoplasma haemofelis, “Candidatus Mycoplasma haemominutum,” and “Candidatus Mycoplasma turicensis.” The way in which these agents are transmitted is largely unknown. Thus, this study aimed to investigate fleas, ticks, and rodents as well as saliva and feces from infected cats for the presence of hemotropic mycoplasmas, to gain insight into potential transmission routes for these agents. DNA was extracted from arthropods and from rodent blood or tissue samples from Switzerland and from salivary and fecal swabs from two experimentally infected and six naturally infected cats. All samples were analyzed with real-time PCR, and some positive samples were confirmed by sequencing. Feline hemotropic mycoplasmas were detected in cat fleas and in a few Ixodes sp. and Rhipicephalus sp. ticks collected from animals but not in ticks collected from vegetation or from rodent samples, although the latter were frequently Mycoplasma coccoides PCR positive. When shedding patterns of feline hemotropic mycoplasmas were investigated, “Ca. Mycoplasma turicensis” DNA was detected in saliva and feces at the early but not at the late phase of infection. M. haemofelis and “Ca. Mycoplasma haemominutum” DNA was not amplified from saliva and feces of naturally infected cats, despite high hemotropic mycoplasma blood loads. Our results suggest that besides an ostensibly indirect transmission by fleas, direct transmission through saliva and feces at the early phase of infection could play a role in the epizootiology of feline hemotropic mycoplasmas. Neither the investigated tick nor the rodent population seems to represent a major reservoir for feline hemotropic mycoplasmas in Switzerland.     

Effect of the MACS technique on rabbit sperm motility

Magnetic-activated cell sorting (MACS) separates apoptotic spermatozoa by the use of annexin V-conjugated nanoparticles which bind to phosphatidylserine that is externalized on the outer leaflet of the sperm plasma membrane. This technique yields two fractions: annexin V-negative (AnV⁻) and annexin V-positive (AnV⁺). The aim of the study was to evaluate the effect of MACS application on the motility parameters of rabbit spermatozoa. Rabbit semen samples collected separately from 4 bucks (I, II, III, and IV) were filtered and separated in a MACS system. The semen samples from a control (untreated) group, AnV⁻ and AnV⁺ fraction were evaluated using CASA system. The experiment was replicated 4 times for each buck. The AnV⁺ sperm had significantly lower concentration than the AnV⁻ fractions and the control samples (P<0.05 for bucks I, II, III, but not IV). We observed that the proportion of apoptotic spermatozoa in the semen of NZW bucks is about 20%. There was no significant difference in the percentage of motile and progressively motile spermatozoa between the AnVfractions and control samples. In conclusion, the MACS technique has no harmful effect on the rabbit sperm concentration and motility.     

Toxoplasma gondii: Evidence for the transmission by semen in dogs

Ten male dogs were distributed into three experimental groups for infection with Toxoplasma gondii: GI - three dogs inoculated with 2.0 × 10⁵ P strais oocysts, GII - three dogs infected with 1.0 × 10⁶ RH strain tachyzoites, and GIII - four controls dogs. Several clinical parameters were evaluated. IFAT was performed to detect anti-T. gondii antibodies. Presence of the parasite in semen was evaluated by PCR and bioassay techniques. Tissue parasitism was examined using bioassays and immunohistochemistry in testicle and epididymis fragments collected after orchiectomy. In semen samples collected from these two groups, the presence of T. gondii was verified by bioassays and PCR. T. gondii was detected by immunohistochemistry in tissues (testicle and epididymis fragments) of all six experimentally infected dogs. The T. gondii-positive seminal samples were used in the artificial insemination (AI) of four female dogs free of toxoplasmic infection. Seven days after AI, all of the female dogs presented serologic conversion (IFAT). Fetal reabsorption occurred in two of the dogs, while the others sustained full-term gestation. Several T. gondii cysts were detected in the brains of four offspring. These results suggest that T. gondii can be sexually transmitted in domestic dogs.