Research Outputs
Citrate-buffered Yamanaka medium allows to produce high-yield bacterial nanocellulose in static culture using Komagataeibacter strains isolated from apple cider vinegar
Bacterial nanocellulose (BNC) is a sustainable, renewable, and eco-friendly nanomaterial, which has gained great attentions in both academic and industrial fields. Two bacterial nanocellulose-producing strains (CVV and CVN) were isolated from apple vinegar sources, presenting high 16S rRNA gene sequence similarities (96%–98%) with Komagataeibacter species. The biofilm was characterized by scanning electron microscopy (SEM), revealing the presence of rod-shaped bacteria intricately embedded in the polymeric matrix composed of nanofibers of bacterial nanocellulose. FTIR spectrum and XRD pattern additionally confirmed the characteristic chemical structure associated with this material. The yields and productivities achieved during 10 days of fermentation were compared with Komagataeibacter xylinus ATCC 53524, resulting in low levels of BNC production. However, a remarkable increase in the BNC yield was achieved for CVV (690% increase) and CVN (750% increase) strains at day 6 of the fermentation upon adding 22 mM citrate buffer into the medium. This effect is mainly attributed to the buffering capacity of the modified Yakamana medium, which allowed to maintain pH close to 4.0 until day 6, though in combination with additional factors including stimulation of the gluconeogenesis pathway and citrate assimilation as a carbon source. In addition, the productivities determined for both isolated strains (0.850 and 0.917 g L−1 d−1) compare favorably to previous works, supporting current efforts to improve fermentation performance in static cultures and the feasibility of scaling-up BNC production in these systems.
Generation of reporter mice for detecting the transcriptional activity of nuclear factor of activated T cells
Nuclear factor of activated T cells (NFAT) is a transcription factor essential for immunological and other biological responses. To develop analyzing system for NFAT activity in vitro and in vivo, we generated reporter mouse lines introduced with NFAT-driven enhanced green fluorescent protein (EGFP) expressing gene construct. Six tandem repeats of −286 to −265 of the human IL2 gene to which NFAT binds in association with its cotranscription factor, activator protein (AP)-1, was conjunct with thymidine kinase minimum promoter and following EGFP coding sequence. Upon introduction of the resulting reporter cassette into C57BL/6 fertilized eggs, the transgenic mice were obtained. Among 7 transgene-positive mice in 110 mice bone, 2 mice showed the designated reporter mouse character. Thus, the EGFP fluorescence of CD4+ and CD8+ T cells in these mice was enhanced by stimulation through CD3 and CD28. Each of phorbol 12-myristate 13-acetate (PMA) and ionomycin (IOM) stimulation weakly but their combined stimulation strongly enhanced EGFP expression. The stimulation-induced EGFP upregulation was also observed following T cell subset differentiation in a different manner. The EGFP induction by PMA + IOM stimulation was more potent than that by CD3/CD28 stimulation in helper T (Th)1, Th2, Th9, and regulatory T cells, while both stimulation conditions displayed the equivalent EGFP induction in Th17 cells. Our NFAT reporter mouse lines are useful for analyzing stimulation-induced transcriptional activation mediated by NFAT in cooperation with AP-1 in T cells.
Notes on the taxonomic status and distribution of some Cylindrotomidae (Diptera, Tipuloidea), with emphasis on Japanese species
A morphological and molecular study of 17 Cylindrotomidae species revealed that the two subspecies of Cylindrotoma distinctissima, the Nearctic C. americana Osten Sacken, 1865, stat. reval. and the Palearctic C. distinctissima (Meigen, 1818), represent separated lineages and consequently are raised to species level. Cylindrotoma japonica Alexander, 1919, syn. nov. and C. distinctissima alpestris Peus, 1952, syn. nov. are now known to be junior synonyms of C. distinctissima. Triogma kuwanai limbinervis Alexander, 1953, syn. nov. and T. nimbipennis Alexander, 1941, syn. nov. are now placed into synonymy under Triogma kuwanai (Alexander, 1913). The Japanese Cylindrotomidae are all redescribed and all available literature and distribution data are summarised. Supplementary descriptions and illustrations for male and female terminalia of Cylindrotoma nigriventris Loew, 1849, Diogma dmitrii Paramonov, 2005, Liogma nodicornis (Osten Sacken, 1865), Phalacrocera replicata (Linnaeus, 1758), P. tipulina Osten Sacken, 1865, and Triogma trisulcata (Schummel, 1829) are provided. The following new distribution records are outlined; Diogma caudata Takahashi, 1960 from Arkhangelsk Oblast, Russia; D. glabrata (Meigen, 1818) from Belarus, Latvia, and Altai Republic, Amur Oblast, Novgorod Oblast, Magadan Oblast, Samara Oblast, and Kuril Islands (Shikotan I and Paramushir I) in Russia; Liogma serraticornis Alexander, 1919 from Khabarovsk Krai, Russia; Phalacrocera replicata from Khabarovsk Krai, Russia; and the presence of Cylindrotoma nigriventris in Altai Republic, Russia is confirmed.
Evolutionary mechanisms underlying the diversification of nuclear factor of activated T cells across vertebrates
The mechanisms of immunity linked to biological evolution are crucial for understanding animal morphogenesis, organogenesis, and biodiversity. The nuclear factor of activated T cells (NFAT) family consists of five members (NFATc1–c4, 5) with different functions in the immune system. However, the evolutionary dynamics of NFATs in vertebrates has not been explored. Herein, we investigated the origin and mechanisms underlying the diversification of NFATs by comparing the gene, transcript and protein sequences, and chromosome information. We defined an ancestral origin of NFATs during the bilaterian development, dated approximately 650 million years ago, where NFAT5 and NFATc1–c4 were derived independently. The conserved parallel evolution of NFATs in multiple species was probably attributed to their innate nature. Conversely, frequent gene duplications and chromosomal rearrangements in the recently evolved taxa have suggested their roles in the adaptive immune evolution. A significant correlation was observed between the chromosome rearrangements with gene duplications and the structural fixation changes in vertebrate NFATs, suggesting their role in NFAT diversification. Remarkably, a conserved gene structure around NFAT genes with vertebrate evolutionary-related breaking points indicated the inheritance of NFATs with their neighboring genes as a unit. The close relationship between NFAT diversification and vertebrate immune evolution was suggested.
MorfometrÃa y diagnóstico molecular de larvas de Anisakis (Nematoda: Ascaridida) en Merluccius gayi (Chordata: Gadiformes) y Dosidicus gigas (Mollusca: Teuthida) en la región del BiobÃo, Chile
Las especies del género Anisakis (Nematoda: Anisakidae) son parásitos marinos con ciclo de vida indirecto. Los crustáceos planctónicos actúan como primeros hospedadores intermediarios, mientras que peces y cefalópodos intervienen como segundos hospedadores intermediarios o paraténicos, finalmente el ciclo se cierra en los cetáceos, mamÃferos marinos que son los principales hospedadores definitivos del género. En el ciclo de vida, las larvas de Anisakis pueden ser ingeridas por el hombre interviniendo como huésped accidental, lo que puede ocasionar anisakiasis, una zoonosis adquirida a través del consumo de peces y cefalópodos crudos o marinados. Estos nemátodos tienen una distribución cosmopolita, sin embargo, su diversidad ha sido escasamente estudiada en el hemisferio Sur. Por tanto, se evaluó la diversidad de las larvas de Anisakis spp., presentes en dos especies de hospederos de Chile, combinando el análisis morfométrico y genético. Para ello, se recolectaron larvas de Anisakis spp. en la cavidad abdominal de la merluza Merluccius gayi y el calamar Dosidicus gigas, procedentes de terminales pesqueros de la región del BiobÃo, Chile. La caracterización morfométrica de las larvas de Anisakis spp., consistió en la medición de la longitud del esófago, ventrÃculo esofágico, cola, longitud total y ancho máximo del cuerpo. Para los análisis genéticos se usó la región molecular nuclear ITS (ITS1-ITS2) y mitocondrial COX2. Los resultados morfométricos revelaron que las larvas extraÃdas de D. gigas son significativamente de mayor longitud que las recolectadas en M. gayi, sugiriendo una alta variabilidad fenotÃpica hospedador-dependiente. Los análisis moleculares y filogenéticos determinaron la presencia de Anisakis pegreffii en ambos hospedadores, sin embargo, demostraron una baja diferenciación genética y diversidad nucleotÃdica entre las secuencias, indicando una escasa variabilidad genética para el conjunto de datos. Este trabajo constituye el primer registro molecular de A. pegreffii en hospedadores intermediario o paraténicos de la costa de Chile.