A bacterial cutinase from Thermobifida fusca, named Tfu_0883, was genetically modified by site-directed mutagenesis to enhance its activity on poly(ethylene terephthalate) (PET). The new mutations tailored the catalytic site for PET, increasing the affinity of cutinase to this hydrophobic substrate and the ability to hydrolyze it. The mutation I218A was designed to create space and the double mutation Q132A/T101A was designed both to create space and to increase hydrophobicity. The activity of the double mutant on the soluble substrate p-nitrophenyl butyrate increased two-fold compared to wild-type cutinase, while on PET both single and double mutants exhibited considerably higher hydrolysis efficiency. The replacement of specific amino acids at the active site was an effective approach for the improvement of the Tfu_0883 cutinase capacity to hydrolyze polyester surfaces. Thus, this study provides valuable insight on how the function and stability of enzymes can be improved by molecular engineering for their application in synthetic fiber biotransformation.

Fish larval development, not least the spectacular process of flatfish metamorphosis, appears to be under complex endocrine control, many aspects of which are still not fully elucidated. In order to obtain data on the functional development of two major endocrine glands, the pituitary and the thyroid, during flatfish metamorphosis, histology, immunohistochemistry and in situ hybridization techniques were applied on larvae of the Atlantic halibut (Hippoglossus hippoglossus), a large, marine flatfish species, from hatching through metamorphosis. The material was obtained from a commercial hatchery. Larval age is defined as day-degrees (D°=accumulated daily temperature from hatching). Sporadic thyroid follicles are first detected in larvae at 142 D°(27 days post-hatch), prior to the completion of yolk sack absorption. Both the number and activity of the follicles increase markedly after yolk sack absorption and continue to do so during subsequent development. The larval triiodothyronine (T 3 ) and thyroxine (T 4 ) content increases, subsequent to yolk absorption, and coincides with the proliferation of thyroid follicles. A second increase of both T 3 and T 4 occurs around the start of metamorphosis and the T 3 content further increases at the metamorphic climax. Overall, the T 3 content is lower than T 4 . The pituitary gland can first be distinguished as a separate organ at the yolk sack stage. During subsequent development, the gland becomes more elongated and differentiates into neurohypophysis (NH), pars distalis (PD) and pars intermedia (PI). The first sporadic endocrine pituitary cells are observed at the yolk sack stage, somatotrophs (growth hormone producing cells) and somatolactotrophs (somatolactin producing cells) are first observed at 121 D°( 23 days post-hatch), and lactotrophs (prolactin producing cells) at 134 D°(25 days post-hatch). Scarce thyrotrophs are evident after detection of the first thyroid follicles (142 D°), but coincident with a phase in which follicle number and activity increase (260 D°). The somatotrophs are clustered in the medium ventral region of the PD, lactotrophs in the anterior part of the PD and somatolactotrophs are scattered in the mid and posterior region of the pituitary. At around 600 D°, coinciding with the start of metamorphosis, somatolactotrophs are restricted to the interdigitating tissue of the NH. During larval development, the pituitary endocrine cells become more numerous. The present data on thyroid development support the notion that thyroid hormones may play a significant role in Atlantic halibut metamorphosis. The time of appearance and the subsequent proliferation of pituitary somatotrophs, lactotrophs, somatolactotrophs and thyrotrophs indicate at which stages of larval development and metamorphosis these endocrine cells may start to play active regulatory roles.

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Abstract Background and Objectives Multicomponent training (MT) combines aerobic, strength, postural, and balance exercises, and may be a promising intervention strategy for dementia. This meta-analysis study aims to systematize evidence concerning the effectiveness of MT in physical fitness, cognition, and functionality on activities of daily living (ADL) in older adults with dementia, and to identify moderation patterns regarding training variables. Research Design and Methods Four databases were systematically searched to locate potential trials through March 2019. A total of 2.312 records were identified and a final set of 17 manuscripts reviewed; of these, 6 satisfied all eligibility criteria. Results Samples sizes ranged from 27 to 170 participants; MT programs lasted between 4 weeks up to 12 months, took place from a daily basis to twice a week, and sessions ranged from 30-60 min. The TESTEX scale was used to analyze the methodological quality, and the funnel plots to assess risk of bias. This meta-analysis revealed that MT interventions benefit older adults with dementia regarding ADL’s performance (ES= 0.313 [0.16 to 0.46]; p<0.01), but the evidence was not sufficiently robust to determine the effectiveness of MT on cognitive function and physical fitness, particularly, on agility. Discussion and Implications MT may be an important non-pharmacological strategy to enhance ADL’s functionality on older adults with dementia. Findings suggest that long-term interventions are more prevailing than high frequency and longer duration exercise sessions. Further evidence is needed for acknowledging its benefits in specific cognitive abilities and physical fitness. This meta-analysis is registered in PROSPERO (no. CRD42020141545).

The immune system of vertebrates consists of several components that partly interact and complement each other. Therefore, the assessment of the overall effectiveness of immune defence requires the simultaneous measurement of different immune components. In this study, we investigated intraspecific variability of innate [i.e. natural antibodies (NAb) and complement] and acquired (i.e. leucocyte profiles) immunity and its relationship with fitness correlates (i.e. blood parasite load and reproductive success in adults and body mass and survival until fledging in nestlings) in the Eurasian kestrel Falco tinnunculus. Immunity differed between nestlings and adults and also between adult males and females. Adult kestrels with higher levels of complement were less parasitised by Haemoproteus, and males with higher values of NAbs showed a higher reproductive success. In nestlings, the H/L ratio was negatively related to body mass. Survival until fledging was predicted by all measured immunological variables of nestlings as well as by their fathers' level of complement. This is the first time that innate immunity is linked to survival in a wild bird. Thus, intraspecific variation in different components of immunity predicts variation in fitness prospects in kestrels, which highlights the importance of measuring innate immune components together with components of the acquired immunity in studies assessing the effectiveness of the immune system in wild animals.

Flatfish metamorphosis is a unique post-embryonic developmental event in which thyroid hormones (THs) drive the development of symmetric pelagic larva into asymmetric benthic juveniles. One of the eyes migrates to join the other eye on the opposite side of the head. Developmental mechanisms at the basis of the acquisition of flatfish anatomical asymmetry remain an open question. Here we demonstrate that an TH responsive asymmetric centre, determined by deiodinase 2 expression, ventrally juxtaposed to the migrating eye in sole (Solea senegalensis) correlates with asymmetric cranial ossification that in turn drives eye migration. Besides skin pigmentation that is asymmetric between dorsal and ventral sides, only the most anterior head region delimited by the eyes becomes asymmetric whereas the remainder of the head and organs therein stay symmetric. Sub-ocular ossification is common to all flatfish analysed to date, so we propose that this newly discovered mechanism is universal and is associated with eye migration in all flatfish.

Thyroid hormones (THs) are essential for embryonic brain development but the genetic mechanisms involved in the action of maternal THs (MTHs) are still largely unknown. As the basis for understanding the underlying genetic mechanisms of MTHs regulation we used an established zebrafish monocarboxylic acid transporter 8 (MCT8) knock-down model and characterised the transcriptome in 25hpf zebrafish embryos. Subsequent mapping of differentially expressed genes using Reactome pathway analysis together with in situ expression analysis and immunohistochemistry revealed the genetic networks and cells under MTHs regulation during zebrafish embryogenesis. We found 4,343 differentially expressed genes and the Reactome pathway analysis revealed that TH is involved in 1681 of these pathways. MTHs regulated the expression of core developmental pathways, such as NOTCH and WNT in a cell specific context. The cellular distribution of neural MTH-target genes demonstrated their cell specific action on neural stem cells and differentiated neuron classes. Taken together our data show that MTHs have a role in zebrafish neurogenesis and suggest they may be involved in cross talk between key pathways in neural development. Given that the observed MCT8 zebrafish knockdown phenotype resembles the symptoms in human patients with Allan-Herndon-Dudley syndrome our data open a window into understanding the genetics of this human congenital condition.