J. Mohan

Effects of hemicastration and castration on foam production and its relationship with fertility in male Japanese quail

Healthy heavy body weight strain of adult male Japanese quail (Coturnix coturnix Japonica) of the same age were used in this study to observe the effect of hemicastration and castration on the frequency of foam discharges from cloacal gland and other related parameters. The quails were housed in individual cages and divided into four groups: control (intact birds), sham-operated control (intact birds with incision), hemicastrated, and castrated groups of birds. Hemicastration and castration were carried out surgically at 10 and 13 weeks of age, respectively. Subsequently, 3 weeks after castration birds were examined for different parameters. Hemicastration caused a significant (P < 0.05) drop in the foam discharge frequency, weight of the foam, and the level of testosterone in the plasma, whereas in castrated group these variables were observed nil or negligible as compared to both of the controls. A suppressive effect of hemicastration was also noticed on the body weight, area of the cloacal gland, as well as percent fertility. Castration induced the drastic regression of the cloacal gland and a significant reduction (P < 0.05) of the body weight was also noted as compared to other groups. Frequencies of foam discharges were twice the number in daytime (06:00-18:00 h) than night, irrespective of the groups. In another study, the effect of characteristics of foam or foam glands on fertility was examined in 77 male birds paired with females. Several characteristics of foam and foam glands were examined for infertility, such as light yellow foam, dark yellow foam, smaller area of foam gland (below 225 mm2), hardness of foam gland and certain unknown factors that contributed 0.23, 0.68, 5.23, 2.27 and 3.64% infertility, respectively. An overall 12.1% birds were found to be infertile. This data indicated that smaller sizes of foam glands might reflect the poorest fertilizing ability of the male birds. From this study, it may be concluded that the cloacal gland may be considered as an external indicator of testicular function of the birds. Characteristics of the cloacal gland or foam or both may be used as a simple and effective tool to predict the fertilizing ability of an individual male in view of the very small semen ejaculate that is very difficult to collect and evaluate for fertility.

Effect of higher dietary vitamin E concentrations on physical and biochemical characteristics of semen in Kadaknath cockerels.

1. This experiment was to investigate the effects of increasing dietary vitamin E on physical and biochemical characteristics of semen in Indian reared Kadaknath (KN) cockerels. DL-α-Tocopherol acetate was used as the source of vitamin E. 2. A total of 135 one-day-old male KN chicks were randomly selected and divided into 9 groups with 15 chicks in each group (3 dietary treatments × 3 replicates). 3. The basal diet contained 15 IU (10 mg) vitamin E/kg and the two experimental diets were supplemented with 150 IU (100 mg) and 300 IU (200 mg) vitamin E/kg (diets T2 and T3, respectively). 4. Physical characteristics in terms of semen volume, sperm concentration, sperm motility and percentage live sperm did not differ significantly, whereas proportion of abnormal and dead spermatozoa were significantly lower and fertility higher in the T2 group. 5. Biochemical characteristics in term of quantities of protein and nitric oxide (NO) did not differ significantly, whereas the quantity of glucose, acid phosphatase (ACP) and vitamin E were significantly higher in the T2 group. 6. In contrast, the quantities of alkaline phosphatase (ALP), glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) were significantly lower in T2 group and higher in the T1 (control) group. 7. From this study it can be concluded that moderate supplementation of dietary vitamin E may be beneficial for physical and biochemical characteristics of semen in Indian reared KN cock.

High doses of dietary zinc induce cytokines, chemokines, and apoptosis in reproductive tissues during regression

In chickens, high levels of dietary zinc cause molting, and the reproductive system undergoes complete remodeling concomitant to feather replacement. In the present study, the expression profiles of cytokines and chemokines were investigated in the ovary and oviduct of control hens and of hens induced to molt by zinc feeding. The zinc-induced feed-intake suppression, the changes in corticosterone levels, the immune cell populations in the reproductive tract, and the apoptosis of reproductive tissues were analyzed. The expression of mRNAs for interleukin-6 (IL-6), interferon-γ (IFN-γ), the avian ortholog of mammalian IL-8 (chCXCLi2), and a chicken MIP-1β-like chemokine (chCCLi2) in the ovary and of mRNAs for IL-1β, IL-6, IFN-γ, transforming growth factor-β2, chCXCLi2, and chCCLi2 in the oviduct were upregulated significantly during zinc-induced molting. A simultaneous feed-intake reduction was observed with higher expression of cytokines and chemokines. The results of the present investigation also suggested that the upregulation of corticosterone was closely associated with the increased expression of cytokines and chemokines. An increase in apoptosis within reproductive tissue during tissue regression was also noted. We had previously observed the upregulation of these cytokines expression in an earlier study (molting by feed withdrawal). However, the pattern and the level of expression were different among these two methods. These findings indicate that cytokines might be a common mediator of tissue regression during molting induced by diverse methods, although the pattern of induction is different. Thus, a high dose of dietary zinc seems to induce reproductive regression via the upregulation of cytokines and chemokines, the suppression of feed intake, and the increase in serum corticosterone, resulting finally in the apoptosis of reproductive tissues.

Protective effects of melatonin in reduction of oxidative damage and immunosuppression induced by aflatoxin B1-contaminated diets in young chicks

In the present study, the ability of aflatoxin B1 (AFB1) to induce endogenous oxidative damages and the possible ameliorating effects of dietary melatonin (MEL) were investigated. Newly hatched broiler chicks (n = 240) were fed aflatoxin‐contaminated diets (0.5 or 1 mg kg−1 diet) with or without MEL (40 mg kg−1 bw) supplementation in diet for 40 days. AFB1 resulted in a significant increase in lipid peroxidation (LPO) in the liver and erythrocytes accompanied with suppression of superoxide dismutase (SOD) and catalase (CAT) enzyme activities of erythrocytes. It also caused a significant reduction in levels of serum proteins and marked elevation in activities of serum transaminases. AFB1 treatment also decreased both humoral and cell‐mediated immune responses in growing chicks. Simultaneous administration of MEL with AFB1 resulted in an obvious improvement in all the tested parameters. Long‐term rather than short‐term administration of MEL was more effective in rendering protection against AFB1‐induced toxicity.

Isolation of a spermatozoa motility inhibiting factor from chicken seminal plasma with antibacterial property

A 78-kDa spermatozoa motility inhibiting factor (SMIF) was purified from chicken (Gallus domesticus) seminal plasma by anion exchange (DE-53) followed by affinity chromatography on concanavalin A-Sepharose. The factor is thermostable and inhibited the spermatozoa motility in a dose dependent manner. In addition, SMIF inhibited the growth of gram negative bacteria, Pasteurella multocida but not gram positive Streptococcus equi. The factor lost its spermatozoa immobilizing property after treatment with trypsin, chymotrypsin or pepsin. The inhibition of SMIF by beta-mercaptoethanol suggest the involvement of disulfide bonds in its activity. Similarly, this property was lost in presence of chicken seminal plasma or incubating SMIF with anti-SMIF antibodies. Evidence is provided for the presence of a high molecular weight protein (> 100 kDa) in chicken seminal plasma that neutralizes the motility inhibiting property of SMIF. No significant decrease in spermatozoa ATP was observed in presence of SMIF suggesting that the loss of spermatozoa motility was due to factors other than depletion in cells energy. Using anti-SMIF antibodies, a cross-reactive protein was identified in the blood, liver and reproductive tissues of chicken and the seminal plasma of cattle and buffalo. However, the cross-reactive protein failed to inhibit chicken spermatozoa motility. The significance of SMIF in chicken seminal plasma is discussed.

Effect of higher levels of dietary selenium on production performance and immune responses in growing Japanese quail

1. The effect of increasing dietary selenium (Se) on production performance and immune responses in growing (0 to 6 weeks) Japanese quail was investigated. 2. One-day-old chicks (240) were randomly selected and divided into 12 groups with 20 chicks in each group (3 dietary treatments × 4 replicates). The basal diet contained 0·2 mg Se/kg and the two experimental diets were supplemented with 0·5 and 1·0 mg Se/kg. 3. Body weight gain, food intake and food conversion ratio and mortality were not affected by Se supplementation. 4. On d 28, antibody responses to inoculated sheep red blood cells were determined. Antibody titres were significantly higher after feeding the two Se-supplemented diets. 5. During week 4, the response to intradermally injected phytohaemagglutinin, an index of the in vivo cell-mediated immune response, was shown to be increased in the groups fed on the Se-supplemented diets. 6. After 6 weeks, the relative weights of the bursa of Fabricius and thymus were greater in the chicks given the Se-supplemented diets but there was no effect on the relative weight of spleen and liver. 7. It is concluded that supplementing the diet with Se has a beneficial effect on immune responses but does not affect production performance in growing Japanese quail.

Development of a new method for sperm RNA purification in the chicken

Currently RNA transcripts are being used as male fertility biomarker for many mammalian species, but research work on chicken is at halt because classical RNA isolation methods are not effective for chicken spermatozoa. Hence, attempts have been made to optimize RNA isolation protocol from chicken sperm by using different methods, and to confirm the presence of sperm-specific transcripts of PRM and PLCZ1. Semen samples were centrifuged at low speed for removing debris like uric acid. Further, 1mL diluted semen was gently placed over 40% PureSperm or 45%/90% Percoll, and centrifuged to remove somatic cells and immature diploid spermatocytes. RNA was isolated from sperm by using RNAzol or TRIzol reagent or RNeasy Micro kit with certain modification, and RNA quantity and quality were evaluated. RNA isolated by using RNAzol or RNeasy Micro Kit yielded good quantity and quality of RNA for downstream applications compared to TRIzol. 40% PureSperm was found effective in removing somatic cells. RT-PCR results showed that sperm RNA samples were negative for CD4 and PTPRC. All the sperm RNA samples were positive for PRM and PLCZ1, markers of sperm RNA.

Cloacal gland foam enhances motility and disaggregation of spermatozoa in Japanese quail (Coturnix japonica).

The adult male Japanese quail produces white foam from the cloacal gland, which is transferred to the female proctodeum during natural mating. The physiological role of foam on quail spermatozoa is still unclear. Therefore, attempts have been made to understand the effect of cloacal foam on motility and metabolism of quail spermatozoa. The profile of various biochemical constitutes in the foam extract was investigated. The addition of foam extract to neat semen completely disaggregated the clumps of spermatozoa leading to vigorous motility. The metabolic rate (MBRT) of the spermatozoa was significantly increased with the addition of foam extract. The foam extract was sub fractionated into seven different fractions by using the molecular cut off devices. Among all the seven sub-fractions from the foam extract, the addition of < 1 KDa sub-fraction contained lactate and has enhanced sperm motility and metabolism. Another fraction (3-10 KDa) has non-protein and non-heparin components which completely disaggregated the clumped quail spermatozoa. However, the remaining fractions did not show any effect on quail spermatozoa. It can be concluded from the present investigation that the lactate present in foam might be a fuel for sperm metabolism and motility. Furthermore, low molecular weight (3-10 KDa) components in the foam may responsible for sperm disaggregation.

Bisphenol A reduces fertilizing ability and motility by compromising mitochondrial function of sperm

Bisphenol A (BPA) acts as an endocrine disruptor, affects animal reproductive success in vivo and affects sperm functions in vitro at environmentally relevant concentrations, leading to reduction in sperm motility and fertilizing ability in fish. The effect of in vitro BPA on avian sperm functions has not been explored. The present study examined the effect of environmentally relevant concentrations of BPA (0 mM, 0.18 mM, 0.37 mM, and 0.74 mM) on sperm functions in chicken in vitro. Sperm were exposed to concentrations of BPA for 30 min and analyzed for motility, fertilizing ability, live sperm percentage, and mitochondrial membrane potential (Δψm). Results showed that BPA at a concentration of 0.74 mM significantly decreased motility, fertilizing ability, live sperm count percentage, and sperm Δψm. Sperm motility was positively correlated with fertility (r = 0.73, p ≤ 0.01), live sperm percentage (r = 0.64, p ≤ 0.01), and high Δψm (r = 0.44, p ≤ 0.01). A dose-dependent and time-dependent effect of BPA was observed on sperm motility at all BPA concentrations. However, sperms fertilizing ability was unaffected in low BPA concentration (0.18 mM and 0.37 mM). A significantly higher percentage of moribund sperm was observed at 0.37 mM and 0.74 mM BPA compared with at 0.18 mM BPA, in the negative control, and in the vehicle control. The present study confirms that environmentally relevant concentrations of BPA are capable of compromising sperm functions, leading to reduction in fertilizing ability of chicken sperm.

Norfloxacin drug induces reproductive toxicity and alters androgen receptor gene expression in testes and cloacal gland of male Japanese quail (Coturnix Japonica)

In an attempt to investigate the reproductive toxicity of norfloxacin in Japanese quail, male quail were given norfloxacin at 20 mg/kg body weight for 14 d. Then reproductive function and androgen receptor (AR) gene expression was examined in treated and control birds. The results of the present study indicate that fertility, cloacal gland area, sperm concentration, and serum testosterone were reduced significantly (p < 0.05) on day 14 in the norfloxacin-treated birds. Upregulation (p < 0.05) of AR mRNA was also seen in the testes on the 14th d of treatment. A trend toward downregulation of AR mRNA was seen in the cloacal gland of norfloxacin-treated birds. Histological observations revealed that norfloxacin induces cellular atrophy in testes and changes in glandular tissue in the cloacal gland. The results of the present study demonstrate that norfloxacin induces testicular toxicity in Japanese quail.