Biocontrol effect of entomopathogenic fungi Metarhizium anisopliae ethyl acetate-derived chemical molecules: An eco-friendly anti-malarial drug and insecticide.

Insect pests represent a major threat to human health and agricultural production. With a current over-dependence on chemical insecticides in the control of insect pests, leading to increased chemical resistance in target organisms, as well as side effects on nontarget organisms, the wider environment, and human health, finding alternative solutions is paramount. The employment of entomopathogenic fungi is one such potential avenue in the pursuit of greener, more target-specific methods of insect pest control. To this end, the present study tested the chemical constituents of Metarhizium anisopliae fungi against the unicellular protozoan malaria parasite Plasmodium falciparum, the insect pests Anopheles stephensi Listen, Spodoptera litura Fabricius, and Tenebrio molitor Linnaeus, as well as the nontarget bioindicator species, Eudrilus eugeniae Kinberg. Fungal crude chemical molecules caused a noticeable anti-plasmodial effect against P. falciparum, with IC50 and IC90 values of 11.53 and 7.65 µg/mL, respectively. The crude chemical molecules caused significant larvicidal activity against insect pests, with LC50 and LC90 values of 49.228-71.846 µg/mL in A. stephensi, 32.542-76.510 µg/mL in S. litura, and 38.503-88.826 µg/mL in T. molitor at 24 h posttreatment. Based on the results of the nontarget bioassay, it was revealed that the fungal-derived crude extract exhibited no histopathological sublethal effects on the earthworm E. eugeniae. LC-MS analysis of M. anisopliae-derived crude metabolites revealed the presence of 10 chemical constituents. Of these chemicals, three major chemical constituents, namely, camphor (15.91%), caprolactam (13.27%), and monobutyl phthalate (19.65%), were highlighted for potential insecticidal and anti-malarial activity. The entomopathogenic fungal-derived crude extracts thus represent promising tools in the control of insect pests and malarial parasites.

Insecticidal efficacy of nanoparticles against Spodoptera frugiperda (J.E. Smith) larvae and their impact in the soil.

The present study aims to evaluate the different nanoparticles (Cu NPs, KI NPs, Ag NPs, Bd NPs, and Gv NPs) against 4th instar Spodoptera frugiperda larvae as well as the microbial toxicity, phytotoxicity, and soil pH. Nanoparticles were tested at three concentrations (1000, 10000, and 100000 ppm) using two methods (food dip and larvae dip) against S. frugiperda larvae. Results (from the larval dip method) showed that among the nanoparticles, the KI NPs caused 63%, 98%, and 98% mortality within 5 days in the treatment of 1000, 10000, and 100000 ppm, respectively. After 24 h post treatment, a 1000 ppm concentration showed 95%, 54%, and 94% germination rates in Metarhizium anisopliae, Beauveria bassiana, and Trichoderma harzianum, respectively. The phytotoxicity evaluation clearly showed that NPs did not affect the morphology of the corn plants after the treatment. The soil nutrient analysis results showed that no effect was observed in soil pH or soil nutrients compared to control treatments. The study clearly showed that nanoparticles are caused toxic effect against S. frugiperda larvae.

Bio-efficacy of insecticidal molecule emodin against dengue, filariasis, and malaria vectors.

Emodin, a compound isolated from Aspergillus terreus, was studied using chromatographic and spectroscopic methods and compound purity (96%) was assessed by TLC. Furthermore, high larvicidal activity against Aedes aegypti-AeA (LC50 6.156 and LC90 12.450 mg/L), Culex quinquefasciatus-CuQ (8.216 and 14.816 mg/L), and Anopheles stephensi-AnS larvae (6.895 and 15.24 mg/L) was recorded. The first isolated fraction (emodin) showed higher pupicidal activity against AeA (15.449 and 20.752 mg/L). Most emodin-treated larvae (ETL) showed variations in acetylcholine esterase, α and ß-carboxylesterases, and phosphatase activities in the 4th instar, indicating the intrinsic differences in their biochemical changes. ETL had numerous altered tissues, including muscle, gastric caeca, hindgut, midgut, nerve ganglia, and midgut epithelium. Acute toxicity of emodin on brine shrimp Artemia nauplii (54.0 and 84.5 mg/L) and the zebrafish Danio rerio (less toxicity observed) was recorded. In docking studies, Emodin interacted well with odorant-binding-proteins of AeA, AnS, and CuQ with docking scores of - 8.89, - 6.53, and - 8.09 kcal mol-1, respectively. Therefore, A. terreus is likely to be effective against mosquito larvicides.

Larvicidal and Antifeedant Effects of Copper Nano-Pesticides against Spodoptera frugiperda (J.E. Smith) and Its Immunological Response.

This study aimed to synthesize and evaluate the efficacy of CuO NPs (copper oxide nanoparticles) with varying test concentrations (10−500 ppm) against larvicidal, antifeedant, immunological, and enzymatic activities against larvae of S. frugiperda at 24 h of treatment. Copper nanoparticles were characterized by using a scanning electron microscope (SEM) and energy dispersive X-ray (EDaX) analysis. The EDaX analysis results clearly show that the synthesized copper nanoparticles contain copper as the main element, and the SEM analysis results show nanoparticle sizes ranging from 29 to 45 nm. The CuO NPs showed remarkable larvicidal activity (97%, 94%, and 81% were observed on the 3rd, 4th, and 5th instar larvae, respectively). The CuO NPs produced high antifeedant activity (98.25%, 98.01%, and 98.42%), which was observed on the 3rd, 4th, and 5th instar larvae, respectively. CuO NPs treatment significantly reduced larval hemocyte levels 24 h after treatment; hemocyte counts and sizes changed in the CuO NPs treatment compared to the control. After 24 h of treatment with CuO NPs, the larval acetylcholinesterase enzyme levels decreased with dose-dependent activity. The present findings conclude that CuO NPs cause remarkable larvicidal antifeedant activity and that CuO NPs are effective, pollution-free green nano-insecticides against S. frugiperda.

Toxicity of Metarhizium flavoviride conidia virulence against Spodoptera litura (Lepidoptera: Noctuidae) and its impact on physiological and biochemical activities.

Insect pests of agricultural crops have establish immunological tolerance against fungal infection caused by pathogens via different humoral and cellular processes. Fungal infection can be prevented by insect antioxidant and detoxifying enzymes, but there is no clear understanding of how they physiologically and biochemically interact. Our study aims to examine the antioxidant and detoxifying enzyme defense systems of the pest insect Spodoptera litura in response to infection by Metarhizium flavoviride. At 48 h following exposure to M. flavoviride, antioxidant enzyme levels were modified, and phenoloxidase and total hemocyte count were decreased significantly. The amount of detoxifying enzymes increased significantly. M. flavoviride appears to directly affect the S. litura immune system and results in decreased immunity. In a bioassay, M. flavoviride was found to be harmful to S. litura larvae in their third and fourth instar stage. M. flavoviride may be an effective tool in the control of S. litura larvae. Such entomopathogenic fungi represent cheaper, pollution free, target specific, promising alternatives to synthetic chemical tools in the for control insect pests.

High-Efficiency Bovine Sperm Sexing Used Magnetic-Activated Cell Sorting by Coupling scFv Antibodies Specific to Y-Chromosome-Bearing Sperm on Magnetic Microbeads.

Sperm sexing technique is favored in the dairy industry. This research focuses on the efficiency of bovine sperm sexing using magnetic-activated cell sorting (MACS) by scFv antibody against Y-chromosome-bearing sperm (Y-scFv) coupled to magnetic microbeads and its effects on kinematic variables, sperm quality, and X/Y-sperm ratio. In this study, the optimal concentration of Y-scFv antibody coupling to the surface of magnetic microbeads was 2-4 mg/mL. PY-microbeads revealed significantly enriched Y-chromosome-bearing sperm (Y-sperm) in the eluted fraction (78.01-81.43%) and X-chromosome-bearing sperm (X-sperm) in the supernatant fraction (79.04-82.65%). The quality of frozen-thawed sexed sperm was analyzed by CASA and imaging flow cytometer, which showed that PY-microbeads did not have a negative effect on X-sperm motility, viability, or acrosome integrity. However, sexed Y-sperm had significantly decreased motility and viability. The X/Y-sperm ratio was determined using an imaging flow cytometer and real-time PCR. PY-microbeads produced sperm with up to 82.65% X-sperm in the X-enriched fraction and up to 81.43% Y-sperm in the Y-enriched fraction. Bovine sperm sexing by PY-microbeads showed high efficiency in separating Y-sperm from X-sperm and acceptable sperm quality. This initial technique is feasible for bovine sperm sexing, which increases the number of heifers in dairy herds while lowering production expenses.

The inhibitory action of plant extracts on the mycelial growth of Ascosphaera apis, the causative agent of chalkbrood disease in Honey bee.

Ascosphaera apis is a fungal pathogen, which causes chalkbrood disease in bees and is threatening beekeeping worldwide. The demand for organic honey for export has lately heightened hence the biological control is the option. This study aimed at the in vitro evaluation of the potency of plant extracts against chalkbrood disease for the possibility of being employed as a biological control strategy. The results showed that the combination of plant extracts from cinnamon with spearmint, cinnamon with lemongrass, cinnamon with geranium, and cinnamon with palmarosa at a concentration of 25% and 12.5% inhibited mycelial growth of A. apis by 100%. This demonstrated the potentiality of combining different plant extracts in controlling this disease. In addition, oregano caused inhibition of up to 100% singly. Conclusively, cinnamon in combination with several extracts has a great potential in curbing this disease while oregano offers an amazing remedy and hence the best formulations should be generated for the beekeeper to utilize.

Improvement of Intramuscular Fat in longissimus Muscle of Finishing Thai Crossbred Black Pigs by Perilla Cake Supplementation in a Low-Lysine Diet.

This study was conducted to find out the effects of perilla cake (PC) supplementation in a low-lysine diet on Thai crossbred finishing pigs' productivity, carcass and meat quality, and fatty acid composition. For six weeks, a total of 21 barrows of finishing pigs were fed with three dietary treatments (T1: basal diet, T2: 2.5 percent PC supplementation in a low-lysine diet, and T3: 4.5 percent PC supplementation in a low-lysine diet). The results show that the intramuscular fat and marbling score was significantly increased by T2 and T3. On the other hand, it was found that the boiling loss and shear force value were significantly decreased by T2 and T3 (p < 0.05). In a low-lysine diet, dietary PC supplementation caused a significant increase in malondialdehyde levels in meat (p < 0.05) compared with the basal diet. It was also shown that alpha-linolenic acid level in backfat and the longissimus thoracis et lumborum muscle was increased considerably by T2 and T3. Therefore, supplementing PC in a low-lysine diet may be an alternative strategy for improving the meat quality of late-phase pigs.

Microsatellite Polymorphism and the Population Structure of Dugongs (Dugong dugon) in Thailand.

The dugong (Dugong dugon) is an endangered species of marine mammals, so knowledge of genetic diversity of these populations is important for conservation planning within different habitats. In this study, six microsatellite markers were used to assess the genetic diversity and population structure of 77 dugongs from skin samples of stranded animals collected from 1994-2019 (69 from Andaman Sea and 8 from the Gulf of Thailand). Our results found that dugongs in the Andaman Sea had higher genetic variation than those in the Gulf of Thailand. Populations in Trang, Satun, and some areas of Krabi had highest diversity compared to other regions of Thailand. Bayesian genetic clustering analysis revealed that dugongs in Thailand consist of five genetic groups. Moreover, dugongs in the middle and lower Andaman Sea presented the greatest gene flow compared to other regions. However, based on calculation of inbreeding coefficients (Fis value = 0.239), dugong populations in the Sea of Thailand are experiencing some levels of inbreeding, and so may warrant special protections. These results provide important information for understanding the genetic status of dugongs that can lead to improved management and conservation of this endangered species.

Association of IFNA16 and TNFRSF19 Polymorphisms with Intramuscular Fat Content and Fatty Acid Composition in Pigs.

Interferon-alpha-16 (IFNA16) and tumor necrosis factor receptor superfamily member 19 (TNFRSF19) are cytokines that may play a role in adipogenesis and fatness. Single nucleotide polymorphisms (SNPs) of the porcine IFNA16 and TNFRSF19 genes were verified and their association with intramuscular fat (IMF) content and fatty acid (FA) composition were evaluated in commercial crossbred pigs. Two non-synonymous SNPs of the porcine IFNA16 c.413G > A and TNFRSF19 c.860G > C loci were detected in commercial crossbred pigs. The porcine IFNA16 c.413G >A polymorphism was significantly associated with stearic acid, total saturated FAs (SFAs), and the ratio of monounsaturated FAs (MUFAs) to SFAs (p < 0.05). Furthermore, the porcine TNFRSF19 c.860G > C polymorphism was found to be significantly associated with IMF content and arachidic acid levels (p < 0.05). The results revealed that porcine IFNA16 and TNFRSF19 polymorphisms are related to IMF content and/or FA composition and affirmed the importance of these cytokine genes as potential candidate genes for lipid deposition and FA composition in the muscle tissue of pigs.

Production of single-chain fragment variable (scFv) antibodies specific to plasma membrane epitopes on bull Y-bearing sperm.

Distinguishing between bull Y- and X-bearing sperm populations is advantageous for techniques using sexed bull semen. The aim of this study was to produce a single-chain fragment variable (scFv) antibody against plasma membrane epitopes on bull Y-bearing sperm. Variable heavy (VH)- and variable light (VL)-region genes generated from a hybridoma cell secreting a specific Y-bearing sperm monoclonal antibody (mAb-1F9) were cloned and expressed. The expected sizes of the DNA bands were ∼350 bp for the VH gene and ∼318 bp for the VL gene. The VH and VL genes were generated and used to construct an scFv gene (∼650 bp), which was expressed in E.coli TG1 cells and produced the corresponding soluble scFv antibody. Compared with the parent mAb-1F9, the scFv antibodies presented a high affinity for Y-bearing sperm and low cross-reactivity with X-bearing sperm. An immunofluorescence analysis confirmed that the scFv antibodies and mAb-1F9 recognize epitopes on the Y-bearing sperm surface. The fluorescence signal was strong on the plasma membrane of Y-bearing sperm but very weak for X-bearing sperm. This study aids the application and production of engineered scFv antibodies specific to Y-bearing sperm to distinguish between Y- and X-bearing sperm populations for techniques involving sexed bull semen.

Evaluation of Native Entomopathogenic Fungi for the Control of Fall Armyworm (Spodoptera frugiperda) in Thailand: A Sustainable Way for Eco-Friendly Agriculture.

Fall armyworm, Spodoptera frugiperda, entered Thailand in late 2018 and has now spread in several regions, with devastating effects in maize and rice production, which are some of the most important cereals in the world. Since then, farmers have utilized the available chemical insecticides to try to control it, but their efforts have been futile. Instead, they have ended up using extraordinary dosages, hence threatening non-target species and other fauna and flora, as well as being costly. In this regard, research has been ongoing, aiming to come up with eco-friendly solutions for this insect. We surveyed and collected various isolates of native entomopathogenic fungi intending to test their efficacy against fall armyworm. Six isolates of entomopathogenic fungi were obtained and identified to Beauveria bassiana based on morphological characteristics and multi-gene phylogenetic analyses. Thereafter, the six isolates of B. bassiana were used to perform efficacy experiments against fall armyworm. Additionally, the glycosyl transferase-like protein 1 (GAS1) gene was analyzed. Consequently, all the isolates showed efficacy against S. frugiperda, with isolate BCMU6 causing up to 91.67% mortality. Further, molecular analysis revealed that all the isolates possess the GAS1 gene, which contributed to their virulence against the insect. This is the first report of utilizing native entomopathogenic B. bassiana to manage S. frugiperda in Thailand, with the revelation of GAS1 as a factor in inducing virulence and cuticle penetration. This study has provided valuable information on the potential development of Beauveria bassiana as an eco-friendly bioinsecticide for the management of fall armyworm in Thailand.

Insecticidal Effect of Zinc Oxide Nanoparticles against Spodoptera frugiperda under Laboratory Conditions.

Fall armyworm Spodoptera frugiperda is a major pest of corn, rice, and sorghum among other crops usually controlled using synthetic or biological insecticides. Currently, the new invention of nanotechnology is taking root in the agricultural industry as an alternative source of pest management that is target-specific, safe, and efficient. This study sought to determine the efficacy of commercial Zinc Oxide (ZnO) nanoparticles (NPs) towards S. frugiperda under laboratory conditions. ZnO NPs were diluted into different concentrations (100-500 ppm), where the baby corn used to feed the S. frugiperda larvae was dipped. The development of the insect feeding on food dipped in ZnO solution was significantly (p < 0.05) affected, and the number of days that the insect took to complete its life cycle had a significant difference compared to the control. There was a significant difference in the adults' emergence in all the concentrations of ZnO NPs compared to the control, with over 90% of the eggs successfully going through the life cycle until adult emergence. Additionally, several body malformations were observed throughout the lifecycle of the insect. Also, the fecundity of the females was greatly affected. The findings of this study suggest the possibility of exploitation of ZnO nanoparticles not only to manage S. frugiperda but to significantly reduce their population in the ecosystem through body deformations, reduced fecundity, reduced oviposition, and hatchability of eggs. It will be a valuable tool in integrated pest management regimens.

Effects of Dietary Perilla Cake Supplementation in Growing Pig on Productive Performance, Meat Quality, and Fatty Acid Profiles.

The objective of this study was to determine the effect of perilla cake (PC) supplementation in a growing pig diet on overall growing performance, meat quality, and fatty acid profile. A total of 24 barrow grower crossbred pigs (Large White × Landrace) × Duroc with an initial average body weight of 26.33 kg were fed with a basal diet supplemented with PC at 0%, 5%, and 10% in (PC0, PC5, and PC10, respectively) for 12 weeks. At the end of the experimental period, pigs were slaughtered to determine carcass traits and meat quality. Back fat, abdominal fat, and longissimus dorsi (LD) muscle were collected to investigate fatty acid composition. The results show that the average daily gain (ADG) in the PC10 significantly increased. However, PC supplementation did not influence carcass traits and meat quality except the color as described by lightness (L*). Dietary PC supplementation significantly increased the α-linolenic acid (ALA, C18:3 cis-9, 12, 15), whereas n6/n3 ratio decreased significantly in all tissues investigated. Thus, it can be concluded that the supplementation of PC in growing pig diet is a potential way to increase the fatty acid composition to that required for healthier meat.

Genetic diversity in a unique population of dugong (Dugong dugon) along the sea coasts of Thailand.

Dugong (Dugong dugon) populations have been shrinking globally, due in large part to habitat fragmentation, degradation and ocean pollution, and today are listed as Vulnerable by the IUCN. Thus, determining genetic diversity in the remaining populations is essential for conservation planning and protection. In this study, measures of inter-simple sequence repeat (ISSR) markers and mtDNA D-loop typing were used to evaluate the genetic diversity of 118 dugongs from skin samples of deceased dugongs collected in Thai waters over a 29-year period. Thirteen ISSR primers revealed that dugongs from the Andaman Sea and Gulf of Thailand exhibited more genetic variation in the first 12 years of the study (1990-2002) compared to the last decade (2009-2019). Dugongs from the Andaman Sea, Trang, Satun and some areas of Krabi province exhibited greater diversity compared to other coastal regions of Thailand. Eleven haplotypes were identified, and when compared to other parts of the world (235 sequences obtained from NCBI), five clades were apparent from a total 353 sequences. Moreover, dugongs from the Andaman Sea were genetically distinct, with a separate haplotype belonging to two clades found only in Thai waters that separated from other groups around 1.2 million years ago. Genetic diversity of dugongs in present times was less than that of past decades, likely due to increased population fragmentation. Because dugongs are difficult to keep and breed in captivity, improved in situ conservation actions are needed to sustain genetically healthy wild populations, and in particular, the specific genetic group found only in the Andaman Sea.

Association of adipocytokine IL-1A and IL-6 genes with intramuscular fat content and fatty acid composition in pigs.

Several adipocytokines are involved in inflammatory and immune responses as well as regulated fat deposition and lipid metabolism in mammals. This study aimed to verify the polymorphisms of the porcine interleukin 1A (IL-1A) and interleukin 6 (IL-6) genes and to assess their association with intramuscular fat (IMF) content and fatty acid (FA) composition in commercial crossbred pigs. Two single nucleotide polymorphisms (SNPs) of the porcine IL-1A g.43722547A>G and IL-6 g.91508173C>T loci were found to be segregating in these crossbred pigs. Furthermore, the porcine IL-1A g.43722547A>G polymorphism was found to be significantly associated with myristic, palmitic, palmitoleic, and eicosadienoic acid levels. Moreover, the porcine IL-6 g.91508173C>T polymorphism was significantly associated with IMF content and homolinolenic acid levels. These results suggest that the polymorphisms of the porcine IL-1A and IL-6 genes correlated with lipid content and FA composition and confirmed the importance of the adipocytokine IL-1A and IL-6 genes as candidate genes for fatty acid composition in the muscles of pigs.

Association of IL-4 and IL-4R Polymorphisms with Litter Size Traits in Pigs.

The interleukin-4 (IL-4) and interleukin-4 receptor (IL-4R) are cytokines that are involved in the immune and reproductive systems. This study aimed to verify the polymorphisms in the porcine IL-4 and IL-4R genes and to assess their effects on litter size traits in commercial pigs. Single nucleotide polymorphisms (SNPs) in the porcine IL-4 and IL-4R genes were genotyped by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. A non-coding SNP of IL-4 g.134993898T > C and a non-synonymous SNP of IL-4R c.1577A > T (amino acid change at position 526, Q526L) were found to be segregating in Landrace sows. The IL-4 g.134993898T > C polymorphism was significantly associated with the number of piglets weaned alive (NWA) trait. The IL-4R c.1577A > T polymorphism was significantly associated with the number born alive (NBA) and NWA traits. Moreover, the accumulation of favorable alleles of these two SNP markers revealed significant associations with the NBA, NWA, and mean weight of piglets at weaning (MWW) traits. These findings indicate that the porcine IL-4 and IL-4R genes may contribute to the reproductive traits of pigs and could be used as candidate genes to improve litter size traits in the pig breeding industry.

Consistency of dark skeletal muscles in Thai native black-bone chickens (Gallus gallus domesticus).

Black-bone chickens (Gallus gallus domesticus) have become economically valuable, particularly in Southeast Asia as a consequence of popular traditional Chinese medical practices. Chickens with whole body organ darkness are considered to have higher value and are, therefore, more often requested. This research study aimed to investigate the darkness in 34 skeletal muscles of 10 Thai black-bone chickens (five males and five females). The evaluation of muscle darkness was done on two levels: (i) a color chart was employed at the macroanatomical level and (ii) by using melanin pigment to evaluate the structure at the microanatomy level. The results revealed that the accumulation of melanin pigment in the muscle tissue was observed in the endomysium, perimysium and epimysium. With respect to the results of the color chart test, iliotibialis lateralis pars preacetabularis, gastrocnemius, fibularis longus and puboischiofemoralis pars medialis showed the highest degree of darkness, while serratus profundus, pectoralis, iliotibialis cranialis, flexor cruris lateralis, and flexor cruris medialis appeared to be the least dark. In addition, we found that the highest and lowest amounts of melanin pigment was noted in the flexor carpi ulnaris and pectoralis (p < 0.05), respectively; however, there was no significant difference (p > 0.05) observed between the sexes. These results reveal that the 34 specified muscles of black-bone chickens showed uneven distribution of darkness due to the differing accumulations of melanin pigments of each muscle.This information may provide background knowledge for a better understanding of melanin accumulation and lead to breeding improvements in Thai black-bone chickens.

Spermatological parameters of immunologically sexed bull semen assessed by imaging flow cytometry, and dairy farm trial.

This study compared the quality parameters of bull semen sexed using an immunological method with those of conventional semen by imaging flow cytometry and applied this semen in dairy farm trials. Semen samples were collected from ten ejaculates from five bulls. Each sample was divided into two treatments: conventional semen (CON) and semen sexed using monoclonal male-specific antibodies combined with the complement system for cytotoxicity reaction (IC-sexed). After obtaining frozen-thawed semen, we used imaging flow cytometry to assess acrosome integrity, sperm sex ratio and viability. Sperm morphology was evaluated using eosin-nigrosin staining. The percentage acrosome integrity did not differ between IC-sexed and CON semen (P = 0.313). The sperm sex ratio showed that the percentage of live X-chromosome-bearing sperm was higher than that of live Y-chromosome-bearing sperm in IC-sexed semen (P = 0.001). IC-sexed semen showed a higher percentage of head and tail defects than did CON semen (P = 0.019). In field trials, 585 cows were subjected randomly to AI with CON or IC-sexed semen. The pregnancy rate of the IC-sexed group did not differ from that of the CON group (P = 0.535). However, IC-sexed semen produced a significantly higher percentage of female calves than did CON semen (P = 0.031). Thus, immunological sexing did not adversely affect the acrosome integrity of sperm. Furthermore, a female calf birth rate of over 74 % can potentially be achieved using IC-sexed semen. These findings could help farmers to replace heifers in their herds.

Distribution of Melanin Pigmentation in 33 Organs of Thai Black-Bone Chickens (Gallus gallus domesticus).

The black-bone chicken (Gallus gallus domesticus) is a breed of chicken that is commonly found in Thailand. This breed is known for having a number of black colored organs. Consumers have been notably attracted to the black-bone chicken breed for the characteristic darkness that is observed in many of its organs. However, the degree of darkness in all organs of the black-bone chicken is still in question. Importantly, there have not yet been any published reports on the distribution of melanin pigment in the organs of the black-bone chicken. This research study aims to examine the distribution of the melanin pigment in 33 organs of the Thai black-bone chicken. Ten black-bone chickens (five male, five female) were included in this study. Thirty-two organs including the brain, spinal cord, sciatic nerve, larynx, trachea, syrinx, lungs, heart, pericardium, aorta, brachial vein, kidney, cloaca, oviduct, testis, gastrocnemius muscle, femur, tongue, esophagus, crop, proventriculus, gizzard, duodenum, jejunum, ileum, cecum, pancreas, liver, gall bladder, omentum, abdominal fat, spleen, and skin were examined in this study. Histological sections taken from tissue samples of each of these organs were studied. The findings revealed that the presence of the melanin pigment was not significantly different (p > 0.005) between male and female specimens. Notably, the liver was the only organ in which the melanin pigment had not accumulated. Consequently, there was not a uniform pattern of melanin pigment accumulation throughout the organs of the chickens. The melanin pigment was present in all of the tissue layers of most organs, while the melanin pigment was found in only specific layers of some of the organs. In conclusion, the distribution of melanin pigmentation in the organs of each of the animals in this study was found to be different. However, in some tissue samples, such as those obtained from the liver, no accumulation of the melanin pigment was observed.