Transcriptional Profiles of Skeletal Muscle Associated With Increasing Severity of White Striping in Commercial Broilers

Development of the white striping (WS) abnormality adversely impacts overall quality of broiler breast meat. Its etiology remains unclear. This study aimed at exploring transcriptional profiles of broiler skeletal muscles exhibiting different WS severity to elucidate molecular mechanisms underlying the development and progression of WS. Total RNA was isolated from pectoralis major of male 7-week-old Ross 308 broilers. The samples were classified as mild (n = 6), moderate (n = 6), or severe (n = 4), based on number and thickness of the white striations on the meat surface. The transcriptome was profiled using a chicken gene expression microarray with one-color hybridization technique. Gene expression patterns of each WS severity level were compared against each other; hence, there were three comparisons: moderate vs. mild (C1), severe vs. moderate (C2), and severe vs. mild (C3). Differentially expressed genes (DEGs) were identified using the combined criteria of false discovery rate 64 0.05 and absolute fold change 651.2. Differential expression of 91, 136, and 294 transcripts were identified in C1, C2, and C3, respectively. There were no DEGs in common among the three comparisons. Based on pathway analysis, the enriched pathways of C1 were related with impaired homeostasis of macronutrients and small biochemical molecules with disrupted Ca2+-related pathways. Decreased abundance of the period circadian regulator suggested the shifted circadian phase when moderate WS developed. The enriched pathways uniquely obtained in C2 were RNA degradation, Ras signaling, cellular senescence, axon guidance, and salivary secretion. The DEGs identified in those pathways might play crucial roles in regulating cellular ion balances and cell-cycle arrest. In C3, the pathways responsible for phosphatidylinositol 3-kinase-Akt signaling, p53 activation, apoptosis, and hypoxia-induced processes were modified. Additionally, pathways associated with a variety of diseases with the DEGs involved in regulation of [Ca2+], collagen formation, microtubule-based motor, and immune response were identified. Eight pathways were common to all three comparisons (i.e., calcium signaling, Ras-associated protein 1 signaling, ubiquitin-mediated proteolysis, vascular smooth muscle contraction, oxytocin signaling, and pathway in cancer). The current findings support the role of intracellular ion imbalance, particularly Ca2+, oxidative stress, and impaired programmed cell death on WS progression

Insights Into Transcriptome Profiles Associated With Wooden Breast Myopathy in Broilers Slaughtered at the Age of 6 or 7 Weeks

open9siThis research was financially supported by Cluster and Program Management, National Science and Technology Development Agency (Thailand; project number P15-50668), and by Office of the Permanent Secretary, Ministry of Higher Education, Science, Research and Innovation (Thailand; P20-50946 and P21-50165).Transcriptomes associated with wooden breast (WB) were characterized in broilers at two different market ages. Breasts (Pectoralis major) were collected, 20-min postmortem, from male Ross 308 broilers slaughtered at 6 and 7 weeks of age. The breasts were classified as “non-WB” or “WB” based on palpation hardness scoring (non-WB = no abnormal hardness, WB = consistently hardened). Total RNA was isolated from 16 samples (n = 3 for 6 week non-WB, n = 3 for 6 week WB; n = 5 for 7 week non-WB, n = 5 for 7 week WB). Transcriptome was profiled using a chicken gene expression microarray with one-color hybridization technique, and compared between non-WB and WB samples of the same age. Among 6 week broilers, 910 transcripts were differentially expressed (DE) (false discovery rate, FDR < 0.05). Pathway analysis underlined metabolisms of glucose and lipids along with gap junctions, tight junction, and focal adhesion (FA) signaling as the top enriched pathways. For the 7 week broilers, 1,195 transcripts were identified (FDR < 0.05) with regulation of actin cytoskeleton, mitogen-activated protein kinase (MAPK) signaling, protein processing in endoplasmic reticulum and FA signaling highlighted as the enriched affected pathways. Absolute transcript levels of eight genes (actinin-1 – ACTN1, integrin-linked kinase – ILK, integrin subunit alpha 8 – ITGA8, integrin subunit beta 5 – ITGB5, protein tyrosine kinase 2 – PTK2, paxillin – PXN, talin 1 – TLN1, and vinculin – VCL) of FA signaling pathway were further elucidated using a droplet digital polymerase chain reaction. The results indicated that, in 6 week broilers, ITGA8 abundance in WB was greater than that of non-WB samples (p < 0.05). Concerning 7 week broilers, greater absolute levels of ACTN1, ILK, ITGA8, and TLN1, accompanied with a reduced ITGB5 were found in WB compared with non-WB (p < 0.05). Transcriptional modification of FA signaling underlined the potential of disrupted cell-cell communication that may incite aberrant molecular events in association with development of WB myopathy.openMalila, Yuwares; Uengwetwanit, Tanaporn; Thanatsang, Krittaporn V.; Arayamethakorn, Sopacha; Srimarut, Yanee; Petracci, Massimiliano; Soglia, Francesca; Rungrassamee, Wanilada; Visessanguan, WonnopMalila, Yuwares; Uengwetwanit, Tanaporn; Thanatsang, Krittaporn V.; Arayamethakorn, Sopacha; Srimarut, Yanee; Petracci, Massimiliano; Soglia, Francesca; Rungrassamee, Wanilada; Visessanguan, Wonno

Differential expression patterns of genes associated with metabolisms, muscle growth and repair in Pectoralis major muscles of fast- and medium-growing chickens

The aim of this study was to investigate the expression of genes related to muscle growth, hypoxia and oxidative stress responses, a multi-substrate serine/threonine-protein kinase (AMPK) and AMPK-related kinases, carbohydrate metabolism, satellite cells activities and fibro- adipogenic progenitors (FAPs) in fast-growing (FG) (n = 30) and medium-growing (MG) chickens (n = 30). Pectoralis major muscles were collected at 7d, 14d, 21d, 28d, 35d and 42d of age. According to their macroscopic features, the samples from FG up to 21d of age were classified as unaffected, while all samples collected at an older age exhibited macroscopic features ascribable to white striping and/or wooden breast abnormalities. In contrast, MG samples did not show any feature associated to muscle disorders. The absolute transcript abundance of 33 target genes was examined by droplet digital polymerase chain reaction. The results showed differential gene expression profiles between FG and MG chickens at different ages. While most genes remained unchanged in MG chickens, the expression patterns of several genes in FG were significantly affected by age. Genes encoding alpha 1, alpha 2, beta 2 and gamma 3 isoforms of AMPK, as well as AMPK-related kinases, were identified as differentially expressed between the two strains. The results support the hypothesis of oxidative stress-induced muscle damage with metabolic alterations in FG chickens. An increased expression of ANXA2, DES, LITAF, MMP14, MYF5 and TGFB1 was observed in FG strain. The results suggest the occurrence of dysregulation of FAP proliferation and differentiation occurring during muscle repair. FAPs could play an important role in defining the proliferation of connective tissue (fibrosis) and deposition of intermuscular adipose tissue which represents distinctive traits of muscle abnormalities. Overall, these findings demonstrate that dysregulated molecular processes associated with myopathic lesions in chickens are strongly influenced by growth rate, and, to some extent, by age

Monitoring of white striping and wooden breast cases and impacts on quality of breast meat collected from commercial broilers ()

Objective This study aimed at investigating white striping (WS) and wooden breast (WB) cases in breast meat collected from commercial broilers. Methods A total of 183 breast samples were collected from male Ross 308 broilers slaughtered at the age of 6 weeks (n = 100) and 7 weeks (n = 83). The breasts were subjected to meat defect inspection, meat quality determination and histology evaluation. Results Of 183, 4 breasts from 6-week-old broilers were classified as non-defective while the others exhibited the WS lesion. Among the 6-week-old birds, the defective samples from the medium size birds (carcass weight ≤2.5 kg) showed mild to moderate WS degree with no altered meat quality. Some of the breasts from the 6-week-old birds with carcass weight above 2.5 kg exhibited WB in accompanied with the WS condition. Besides of a reduction of protein content, increases in collagen matter and pH values in the defective samples (p<0.05), no other impaired quality indices were detected within this group. All 7-week-old broilers yielded carcasses weighing above 2.5 kg and showed abnormal characteristics with progressive severity. The breasts affected with severe WS and WB showed the greatest cook loss, hardness, springiness and chewiness (p<0.05). Development of WB induced significantly increased drip loss in the samples (p<0.05). Histology indicated necrotic events in the defective myofibers. Based on logistic regression, increasing percent breast weight by one unit enhanced the chance of WS and WB development with advanced severity by 50.9% and 61.0%, respectively. Delayed slaughter age from 6 to 7 weeks increased the likelihood of obtaining increased WS severity by 56.3%. Conclusion Cases of WS and WB defects in Southeast Asia have been revealed. Despite few cases of the severe WS and WB, such abnormal conditions significantly impaired technological properties and nutritional quality of broiler breasts

Oxidative stress responses in Escherichia coli and Pseudomonas aeruginosa

Aerobic respiration generates unavoidable reactive oxygen species (ROS, detrimental by-products that damage biological molecules. The accumulation of ROS beyond the cellular capacity for detoxification results in oxidative stress. Bacteria have evolved transcription factors that sense ROS and transduce signals into differential gene expression. This study focuses on oxidative stress responses in Escherichia coli, an enteric bacterium and Pseudomonas aeruginosa, an important opportunistic human pathogen. E. coli activates the response to paraquat-induced superoxide stress through the SoxRS system. This study showed that transcription of pgi, a SoxRS regulated gene, and the corresponding phosphoglucose isomerase activity were induced in a SoxRS-dependent manner. A pgi null mutant was hypersensitive to paraquat when grown in rich medium with glucose. We provide evidence that pgi plays a role in maintaining NADPH for cells under oxidative stress by providing glucose-6-phosphate as a substrate for glucose-6-phosphate dehydrogenase. The opportunistic human pathogen P. aeruginosa has evolved under self-generated redox-cycling compounds, in addition to physiological and host-derived sources of oxidative stress. In an effort to identify novel redox sensor and effector proteins, we characterized the pqrCBAR genes. The mutation of the pqrCBAR genes resulted in hypersensitivity to oxidants, and pqrCBA form an operon under negative transcriptional regulation by PqrR. PqrR, a member of MarR family transcription regulators, has 4 conserved cysteines. Purified PqrR was found to exist as a dimer in solution, contain iron, and display characteristic UV spectra common to proteins with iron-sulfur clusters. DNAseI protection assay showed that PqrR bound in vitro at two distinct sites of the pqrA-pqrR intergenic region. PqrR binding activity was reduced by exposure to air, and reconstituted by DTT treatment. The site-directed mutagenesis of PqrR revealed that 3 conserved cysteine residues were essential for redox-sensing activity. To identify other novel antioxidant mechanisms in P. aeruginosa, we screened 15,003 of random transposon insertion mutants, and confirmed 15 mutants for paraquat sensitive phenotype

The PqrR Transcriptional Repressor of Pseudomonas aeruginosa Transduces Redox Signals via an Iron-Containing Prosthetic Group▿ †

Inducible defenses against oxidative stress are coordinated by redox-sensitive transcription factors that transduce oxidative damage into differential gene expression. The opportunistic human pathogen Pseudomonas aeruginosa has evolved under physiological and host-derived sources of oxidative stress. Previous work showed that the pqrABC and pqrR genes of P. aeruginosa, all lacking known functions, were induced by treatment of three different isolates of P. aeruginosa with paraquat (PQ), a superoxide-producing agent. Insertional mutation of the pqrABCR genes resulted in hypersensitive phenotypes to a variety of oxidants, although the hypersensitivity to PQ was marginal. Mutation of pqrR and complementation assays showed that PqrR regulated the pqrABC genes in response to PQ. PqrR, a member of the MarR family of transcriptional regulators, contains a C-terminal region with four conserved cysteines, which suggested redox-regulated transcriptional activity. Purified PqrR bound to two discrete sites at the pqrA and pqrR regulatory regions. The in vitro DNA binding activity of PqrR was decreased by exposure to air and reconstituted by treatment with dl-dithiothreitol. Elemental analysis and preliminary electron paramagnetic resonance experiments showed that PqrR contains iron. Interestingly, site-directed mutagenesis of C-terminal cysteines demonstrated that the four conserved cysteine residues are essential for in vivo redox sensing by PqrR

Improvement of Moist Heat Resistance of Ascorbic Acid through Encapsulation in Egg Yolk&ndash;Chitosan Composite: Application for Production of Highly Nutritious Shrimp Feed Pellets

Egg yolk (EY) is an excellent supplement for aquatic animals and has good technofunctionality. Ascorbic acid (AA) is a potent bioactive substance and is essentially added to shrimp feed; however, it is drastically lost in both feed processing and in rearing environments. In this study, AA was microencapsulated in an EY&ndash;chitosan (CS) composite. The encapsulated vitamin was then mixed into a shrimp feed mixture to form pelleted feed via twin-screw extrusion. The effects of the EY/AA ratio and the amount of CS on moist heat resistance, production yield, encapsulation efficiency (EE), and morphology of microcapsules were investigated. The molecular interaction of the microcapsule components was analyzed by FTIR. The size and size distribution of the microcapsules were determined using a laser diffraction analyzer. The microstructure was evaluated by SEM. The physical properties of the microcapsule-fortified pelleted feed were determined. The AA retention at each step of feed processing and during exposure to seawater was evaluated. The results showed that the microcapsules had a spherical shape with an average diameter of ~6.0 &mu;m. Decreasing the EY/AA ratio significantly improved the production yield, EE, and morphology of the microcapsules. EY proved to be the key component for moist heat resistance, while CS majorly improved the production yield, EE, and morphology of the microcapsules. The microcapsules showed no adverse impact on feed properties. The loss of AA in food processing and seawater was remarkably improved. The final content of the encapsulated AA remaining in shrimp feed was 16-fold higher than that of the unencapsulated AA

Bacterial population in intestines of the black tiger shrimp (Penaeus monodon) under different growth stages.

Intestinal bacterial communities in aquaculture have been drawn to attention due to potential benefit to their hosts. To identify core intestinal bacteria in the black tiger shrimp (Penaeus monodon), bacterial populations of disease-free shrimp were characterized from intestines of four developmental stages (15-day-old post larvae (PL15), 1- (J1), 2- (J2), and 3-month-old (J3) juveniles) using pyrosequencing, real-time PCR and denaturing gradient gel electrophoresis (DGGE) approaches. A total of 25,121 pyrosequencing reads (reading length = 442±24 bases) were obtained, which were categorized by barcode for PL15 (7,045 sequences), J1 (3,055 sequences), J2 (13,130 sequences) and J3 (1,890 sequences). Bacteria in the phyla Bacteroides, Firmicutes and Proteobacteria were found in intestines at all four growth stages. There were 88, 14, 27, and 20 bacterial genera associated with the intestinal tract of PL15, J1, J2 and J3, respectively. Pyrosequencing analysis revealed that Proteobacteria (class Gammaproteobacteria) was a dominant bacteria group with a relative abundance of 89% for PL15 and 99% for J1, J2 and J3. Real-time PCR assay also confirmed that Gammaproteobacteria had the highest relative abundance in intestines from all growth stages. Intestinal bacterial communities from the three juvenile stages were more similar to each other than that of the PL shrimp based on PCA analyses of pyrosequencing results and their DGGE profiles. This study provides descriptive bacterial communities associated to the black tiger shrimp intestines during these growth development stages in rearing facilities

Characterization of Intestinal Bacteria in Wild and Domesticated Adult Black Tiger Shrimp (<i>Penaeus monodon</i>)

<div><p>The black tiger shrimp (<i>Penaeus monodon</i>) is a marine crustacean of economic importance in the world market. To ensure sustainability of the shrimp industry, production capacity and disease outbreak prevention must be improved. Understanding healthy microbial balance inside the shrimp intestine can provide an initial step toward better farming practice and probiotic applications. In this study, we employed a barcode pyrosequencing analysis of V3-4 regions of 16S rRNA genes to examine intestinal bacteria communities in wild-caught and domesticated <i>P. monodon</i> broodstock. Shrimp faeces were removed from intestines prior to further analysis in attempt to identify mucosal bacterial population. Five phyla, <i>Actinobacteria</i>, <i>Fusobacteria</i>, <i>Proteobacteria</i>, <i>Firmicutes</i> and <i>Bacteroidetes,</i> were found in all shrimp from both wild and domesticated environments. The operational taxonomic unit (OTU) was assigned at 97% sequence identity, and our pyrosequencing results identified 18 OTUs commonly found in both groups. Sequences of the shared OTUs were similar to bacteria in three phyla, namely <i>i) Proteobacteria</i> (<i>Vibrio</i>, <i>Photobacterium</i>, <i>Novosphingobium</i>, <i>Pseudomonas</i>, <i>Sphingomonas</i> and <i>Undibacterium</i>), <i>ii) Firmicutes</i> (<i>Fusibacter</i>), and <i>iii) Bacteroidetes</i> (<i>Cloacibacterium</i>). The shared bacterial members in <i>P. monodon</i> from two different habitats provide evidence that the internal environments within the host shrimp also exerts selective pressure on bacterial members. Intestinal bacterial profiles were compared using denaturing gradient gel electrophoresis (DGGE). The sequences from DGGE bands were similar to those of <i>Vibrio</i> and <i>Photobacterium</i> in all shrimp, consistent with pyrosequencing results. This work provides the first comprehensive report on bacterial populations in the intestine of adult black tiger shrimp and reveals some similar bacterial members between the intestine of wild-caught and domesticated shrimp.</p></div