Molecular characterization of a teleost-specific toll-like receptor 22 (tlr22) gene from yellow catfish (Pelteobagrus fulvidraco) and its transcriptional change in response to poly I:C and Aeromonas hydrophila stimuli.

Toll-like receptors (TLRs) are a class of pattern recognition receptors (PRRs) that can recognize pathogen-associated molecular patterns (PMPs) and play important roles in the innate immune system in vertebrates. In this study, we identified a teleost-specific tlr22 gene from yellow catfish (Pelteobagrus fulvidraco) and its immune roles in response to different pathogens were also determined. The open reading frame (ORF) of the tlr22 was 2892 bp in length, encoding a protein of 963 amino acids. Multiple protein sequences alignment, secondary and three-dimensional structure analyses revealed that TLR22 is highly conserved among different fish species. Phylogenetic analysis showed that the phylogenetic topology was divided into six families of TLR1, TLR3, TLR4, TLR5, TLR7 and TLR11, and TLR22 subfamily was clustered into TLR11 family. Meanwhile, synteny and gene structure comparisons revealed functional and evolutionary conservation of the tlr22 gene in teleosts. Furthermore, tlr22 gene was shown to be widely expressed in detected tissues except barbel and eye, with highest expression level in liver. The transcription of tlr22 was significantly increased in spleen, kidney, liver and gill tissues at different timepoints after Poly I:C infection, suggesting TLR22 plays critical roles in defensing virus invasion. Similarly, the transcription of tlr22 was also dramatically up-regulated in spleen, kidney and gill tissues with different patterns after Aeromonas hydrophila infection, indicating that TLR22 is also involved in resisting bacteria invasion. Our findings will provide a solid basis for the investigation the immune functions of tlr22 gene in teleosts, as well as provide useful information for disease control and treatment for yellow catfish.

Toll-like Receptor 3 in the Hybrid Yellow Catfish (Pelteobagrus fulvidraco ♀ × P. vachelli ♂): Protein Structure, Evolution and Immune Response to Exogenous Aeromonas hydrophila and Poly (I:C) Stimuli.

As a major mediator of cellular response to viral infection in mammals, Toll-like receptor 3 (TLR3) was proved to respond to double-stranded RNA (dsRNA). However, the molecular mechanism by which TLR3 functions in the viral infection response in teleosts remains to be investigated. In this study, the Toll-like receptor 3 gene of the hybrid yellow catfish was identified and characterized by comparative genomics. Furthermore, multiple sequence alignment, genomic synteny and phylogenetic analysis suggested that the homologous TLR3 genes were unique to teleosts. Gene structure analysis showed that five exons and four introns were common components of TLR3s in the 12 examined species, and interestingly the third exon in teleosts was the same length of 194 bp. Genomic synteny analysis indicated that TLR3s were highly conserved in various teleosts, with similar organizations of gene arrangement. De novo predictions showed that TLR3s were horseshoe-shaped in multiple taxa except for avian (with a round-shaped structure). Phylogenetic topology showed that the evolution of TLR3 was consistent with the evolution of the studied species. Selection analysis showed that the evolution rates of TLR3 proteins were usually higher than those of TLR3-TIR domains, indicating that the latter were more conserved. Tissue distribution analysis showed that TLR3s were widely distributed in the 12 tested tissues, with the highest transcriptions in liver and intestine. In addition, the transcription levels of TLR3 were significantly increased in immune-related tissues after infection of exogenous Aeromonas hydrophila and poly (I:C). Molecular docking showed that TLR3 in teleosts forms a complex with poly (I:C). In summary, our present results suggest that TLR3 is a pattern recognition receptor (PRR) gene in the immune response to pathogen infections in hybrid yellow catfish.

Transcriptome characterization and SSR discovery in the giant spiny frog Quasipaa spinosa.

The giant spiny frog Quasipaa spinosa (Amphibia: Ranidae) is a large unique frog species found mainly in southern China with a low amount of fat and high protein, and it has become one of the most important aquaculture animal species in China. To better understand its genetic background and screen potential molecular markers for artificial breeding and species conservation, we constructed an expression profile of Q. spinosa with high-throughput RNA sequencing and acquired potential SSR markers. Approximately 81.7 Gb of data and 93,887 unigenes were generated. The transcriptome contains 2085 (80.7 %) complete BUSCOs, suggesting that our assembly methods were effective and accurate.These unigenes were functionally classified using 7 functional databases, yielding 17,482 Pfam-, 12,752 Sting-, 17,526 KEGG-, 24,341 Swiss-Prot-, 28,604 Nr-, 16,287 GO- and 12,752 COG-annotated unigenes. Among several amphibian species, Q. spinosa unigenes had the highest number of hits to Xenopus tropicalis (35.25 %), followed by Xenopus laevis (12.68 %). 1417 unigenes were assigned to the immune system. In addition, a total of 33,019 candidate SSR markers were identified from the constructed library. Further tests with 20 loci and 118 large-scale breeding specimens gathered from four culture farms in China showed that 15 (75 %) loci were polymorphic, with the number of alleles per locus varying from 3 to 9 (mean of 4.3). The PIC values for the SSR markers ranged from 0.19 to 0.82, with an average value of 0.43, indicating moderate polymorphism in Q. spinosa. The transcriptomic profile and SSR repertoire obtained in the present study will facilitate population genetic studies and the selective breeding of amphibian species.

Which Seat Facilitates the Detection of Off-Seat Behaviours? An Inattentional Blindness Test on Location Effect in the Classroom.

Off-seat behaviour refers to students leaving their seats and walking out of a classroom without the teacher noticing. This behaviour occurs in special education for students with certain special needs, which would lead to serious safety problems. This study carried out an inattentional blindness test to explore whether the location of seats in classrooms would impact teachers' detection rate regarding off-seat behaviours. The participants were 126 pre-service teachers (Mage = 18.72 ± 0.723; 92% female) who were invited to perform the primary task of counting students raising their hands up whilst the disappearance of one of the students was introduced as an unexpected occurrence. The results show that peripheral seats were more detectable than the central ones for the teachers to notice the "missing student." Meanwhile, the left and below oriented seats were more likely to be ignored compared to those that were right and upper oriented. These results suggest the existence of a location effect in the classroom that is associated with teachers' attention regarding off-seat behaviour. This study has implications for classroom management in terms of arranging students' seats appropriately to assist in increasing teachers' identification of this hazard.

The effect of emotional valence and age of faces on adults and children's inattentional blindness.

Inattentional blindness (IB) refers to the nature of an individual being unaware of an unexpected stimulus when focusing on an attentional task. Investigation into IB provides an innovative approach for the research of attentional bias that is connected with an individual's immediate attentional capture. This study explored the effect of emotional valence and age of faces on the IB rates of children and adults. We employed a between-subject experimental design with a 2 (age of participants: child, adult) × 2 (age of face: child face, adult face) × 2 (emotional valence: positive, negative) method and used the classic IB task of cross judgment. The participants included 79 adults (54.63 ± 8.192 years old) and 78 children (10.62 ± 2.32 years old). The results showed that: (1) The IB rates of the two groups differed significantly. Children performed better than adults in detecting the faces appearing unexpectedly. (2) The effect of emotional valence only emerged on adult participants' IB. Adults were significantly more likely to detect the positive-emotional faces than the negative-emotional ones. (3) The age of a face was not a significant factor to predict participants' IB. Neither children nor adults showed a preference for the faces of a specific age (e.g., their own age). These findings revealed the distinct characteristics of attentional capture of children and adults when confronting unexpected facial stimuli.

Effects of losartan on the mobilization of endothelial progenitor cells and improvement of endothelial function.

Through investigating the effect of the angiotensin receptor blocker (ARB) losartan on the number of endothelial progenitor cells (EPCs) and blood flow-mediated endothelium-dependent function (FMD) in the peripheral blood of patients with coronary heart disease (CHD), we found that FMD was improved and the number of circulating EPCs increased in the ARB treatment group (P <0.05). In addition, the increase in the number of EPCs was positively correlated with the improvement of FMD in the ARB treatment group (r = 0.52, P <0.01). These findings suggest that losartan may mobilize EPCs in the peripheral blood, improving endothelial function in CHD.

Development of a rapid and specific loop-mediated isothermal amplification detection method that targets Marek's disease virus meq gene.

A rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) method was developed and evaluated for the detection of Marek's disease virus (MDV) by amplification of conserved MDV meq gene sequences. LAMP is an innovative technique that allows the rapid detection of targeted nucleic acid sequences under isothermal conditions without the need for complex instrumentation. In this study, meq gene sequences were amplified successfully from different MDV strains by LAMP within 60min and no cross-reactivity was observed in a panel of related viruses that were associated with diseases of chickens. The detection limit of LAMP was 3.2 copies/million cells compared with 320 copies/million cells required for conventional PCR. Positive detection rates were assessed using either LAMP or PCR by examination of feather follicles that were collected from chickens infected experimentally with either strain J-1 (n=20) or strain Md5 (n=17), In addition to these samples, three isolates that were suspected to have been infected in the clinic were also tested. Results showed that the positive detection rate for LAMP was 95% (38/40), compared with 87.5% (35/40) and 90% (38/40) for strains J-1 and Md5 by PCR, respectively. These results indicated that the LAMP assay was more sensitive, rapid and specific than conventional PCR for the detection of MDV. This easy-to-perform technique will be useful for the detection of MDV and will aid in the establishment of disease control protocols.