Atacama desert is a source of new Micromonospora strains: description of Micromonospora sicca sp. nov.

Several strains were isolated from subsurface soil of the Atacama Desert and were previously assigned to the Micromonospora genus. A polyphasic study was designed to determine the taxonomic affiliation of isolates 4G51T, 4G53, and 4G57. All the strains showed chemotaxonomic properties in line with their classification in the genus Micromonospora, including meso-diaminopimelic acid in the cell wall peptidoglycan, MK-9(H4) as major respiratory quinone, iso-C15:0 and iso-C16:0 as major fatty acids and diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylinositol as major polar lipids. The 16S rRNA gene sequences of strains 4G51T, 4G53, and 4G57 showed the highest similarity (97.9 %) with the type strain of Micromonospora costi CS1-12T, forming an independent branch in the phylogenetic gene tree. Their independent position was confirmed with genome phylogenies, being most closely related to the type strain of Micromonospora kangleipakensis. Digital DNA-DNA hybridization and average nucleotide identity analyses between the isolates and their closest phylogenomic neighbours confirmed that they should be assigned to a new species within the genus Micromonospora for which the name Micromonospora sicca sp. nov. (4G51T=PCM 3031T=LMG 30756T) is proposed.

Flavobacterium facile sp. nov., isolated from water system of Atlantic salmon (Salmo salar) fry cultured in Chile.

Strain T-12T, an orange, Gram-stain-negative, non-motile, rod-shaped strain, was isolated in November 2013 from water samples collected from an Atlantic salmon (Salmo salar) fry culturing system at a fish farm in Chile. Phylogenetic analysis based on 16S rRNA sequences (1394 bp) revealed that strain T-12T belonged to the genus Flavobacterium, showing close relationships to Flavobacterium bernardetii F-372T (99.48 %) and Flavobacterium terrigena DS-20T (98.50 %). The genome size of strain T-12T was 3.28 Mb, with a G+C content of 31.1 mol%. Genome comparisons aligned strain T-12T with Flavobacterium bernardetii F-372T (GCA_011305415) and Flavobacterium terrigena DSM 17934T (GCA_900108955). The highest digital DNA-DNA hybridization (dDDH) values were 42.6 % with F. bernardetii F-372T (GCA_011305415) and 33.9 % with F. terrigena DSM 17934T (GCA_900108955). Pairwise average nucleotide identity (ANI) calculations were below the species cutoff, with the best results with F. bernardetii F-372T being: ANIb, 90.33 %; ANIm, 91.85 %; and TETRA, 0.997 %. These dDDH and ANI results confirm that strain T-12T represents a new species. The major fatty acids were iso-C15 : 0 and C15 : 1ω6с. Detected polar lipids included phospholipids (n=2), aminophospholipid (n=1), aminolipid (n=1) and unidentified lipids (n=2). The predominant respiratory quinone was menaquinone MK7 (80 %) followed by MK-6 (20 %). Phenotypic, chemotaxonomic, and genomic data support the classification of strain T-12T (=CECT 30410T=RGM 3222T) as representing a novel species of Flavobacterium, for which the name Flavobacterium facile sp. nov. is proposed.

Lipidomics profile and volatile compounds of squids (Illex argentinus, Ommastrephes Bartram and Dosidicus gigas) in the Argentine, North Pacific Ocean, Equator and Peru by UPLC-triple TOF-MS and HS-SPME-GC-O-MS.

Comprehensive lipid and volatile compound analyses were performed with squids collected from four varied geographical locations to discriminate the regional characteristics. A total of 1442 lipid molecules and 110 volatiles were detected in the squid muscle samples. There were significant differences in the lipid profiles between Argentine squid (Illex argentinus, AGT), North Pacific Ocean squid (Ommastrephes Bartram, NPO), Equatorial squid (Dosidicus gigas, EQ), and Peruvian squid (Dosidicus gigas, PR) muscle. Phosphatidylcholines (14.64%), triacylglycerols (12.42%), and ceramides (10.97%) were the main lipid components. The contents of polyunsaturated fatty acid in phospholipids and in glycerolipids were 30.35-52.05% and 18.11-25.15%, respectively. The volatiles in squids exhibited significant regional variation; 1-pentanol and 1-octanol, 2-ethyl-1-hexanol and terpinen-4-ol, 2,7-ethyl-1-hexanol, 3-methy-1-butanol and 2-propyl-1-pentanol were identified as characteristic flavor compounds in AGT, NPO, EQ, and PR, respectively. Sphingomyelin, phosphatidylserine, phosphatidylethanolamine, and ceramide were strongly correlated with volatiles in squid muscle. Our study is a reference for the lipid nutritional value and flavor compounds of squids.

Microbacterium plantarum sp. nov. and Microbacterium thalli sp. nov., two endophytic metal-resistant bacteria isolated from Sphaeralcea angustifolia (Cav.) G. Don and Prosopis laevigata (Humb. et Bonpl. ex Willd) M.C. Johnston.

Four Gram-positive, aerobic, catalase- and oxidase-negative, rod-shaped, motile endophytic bacterial strains, designated NM3R9T, NE1TT3, NE2TL11 and NE2HP2T, were isolated from the inner tissues (leaf and stem) of Sphaeralcea angustifolia and roots of Prosopis laevigata. They were characterized using a polyphasic approach, which revealed that they represent two novel Microbacterium species. Phylogenetic analysis based on 16S rRNA gene sequencing showed that the species closest to NE2HP2T was Microbacterium arborescens DSM 20754T (99.6 %) and that closest to NM3R9T, NE2TL11 and NE2TT3 was Microbacterium oleivorans NBRC 103075T (97.4 %). The whole-genome average nucleotide identity value between strain NM3R9T and Microbacterium imperiale DSM 20530T was 90.91 %, and that between strain NE2HP2T and M. arborecens DSM 20754T was 91.03 %. Digital DNA-DNA hybridization showed values of less than 70 % with the type strains of related species. The polar lipids present in both strains included diphosphatidylglycerol, phosphatidylglycerol, glycolipids and unidentified lipids, whereas the major fatty acids included anteiso-C15 : 0, anteiso-C17 : 0, iso-C16 : 0 and C16 : 0. Whole-cell sugars included mannose, rhamnose and galactose. Strains NM3R9T and NE2HP2T showed physiological characteristics different from those present in closely related Microbacterium species. According to the taxonomic analysis, both strains belong to two novel species. The name Microbacterium plantarum sp. nov. is proposed for strain NE2HP2T (=LMG 30875T=CCBAU 101117T) and Microbacterium thalli sp. nov. for strains NM3R9T (=LMG 30873T=CCBAU 101116T), NE1TT3 (=CCBAU 101114) and NE2TL11 (=CCBAU 101115).

Rice Bran Lipidome Identifies Novel Phospholipids, Glycolipids, and Oxylipins with Roles in Lipid Metabolism of Hypercholesterolemic Children.

SCOPE: The purpose of the study is to characterize the chemical diversity in rice bran (RB) lipidome and determines whether daily RB consumption for 4 weeks may modulate plasma lipid profiles in children. METHODS AND RESULTS: Untargeted and targeted lipidomics via ultra-performance liquid chromatography coupled with high-resolution tandem mass spectrometry (UPLC-MS/MS) are applied to identify bioactive RB lipids from a collection of 17 rice varieties. To determine the impact of RB (Calrose-USA variety) supplementation on plasma lipid profile, a secondary analysis of plasma lipidome is conducted on data recorded in a clinical study (NCT01911390, n = 18 moderately hypercholesterolemic children) before and after 4 weeks of dietary intervention with a control or RB supplemented (15 g day-1 ) snack. Untargeted lipidomic reveals 118 lipids as the core of lipidome across all varieties among which phospholipids are abundant and oxylipins present. Phytoprostanes and phytofurans are quantified and characterized. Lipidome analysis of the children plasma following RB consumption reveals the presence of polar lipids and oxylipins alongside putative modulations in endocannabinoids associated with RB consumption. CONCLUSION: The investigation of novel polar lipids, oxylipins, phytoprostanes, and phytofurans in RB extracts provides support for new health-promoting properties interesting for people at risk for cardiometabolic disease.

Fisiología respiratoria: fisiología del surfactante pulmonar / Physiology of pulmonary surfactant

El proceso de respiración y el intercambio gaseoso requiere la interacción de variadas fuerzas en los distintos tejidos y órganos involucrados. La tensión superficial a nivel alveolar provocaría colapso de dichas estructuras de no ser por las características del surfactante que lo recubre. Revisaremos en este articulo la fisiología involucrada en su estructura física, producción y efectos pulmonares.

The process of breathing and gas exchange requires the interaction of various forces in the different tissues and organs involved. The surface tension at the alveolus would cause collapse of these structures without of the surfactant that covers it. We will review in this article the physiology involved in its physical structure, production, and pulmonary effects.

Vibrio fluminensis sp. nov. isolated from the skin of Southern Atlantic sharpnose-pufferfish (Canthigaster figueiredoi).

This manuscript provides the description of the bacterial strain A621T characterized by Gram negative motile rods, presenting green circular colonies on TCBS. It was obtained from the skin of the sharpnose pufferfish Canthigaster figueredoi (Tetraodontidae Family), collected in Arraial do Cabo, located in the Rio de Janeiro region, Brazil. Optimum growth occurs at 20-28 °C in the presence of 3% NaCl. The Genome sequence of the novel isolate consisted of 4.224 Mb, 4431 coding genes and G + C content of 44.5%. Genomic taxonomy analysis based on average amino acid (AAI), genome-to-genome-distance (GGDH) and phylogenetic reconstruction placed (A621T= CBAS 741T = CAIM 1945T = CCMR 150T) into a new species of the genus Vibrio (Vibrio fluminensis sp. nov). The genome of the novel species contains four gene clusters (~ 56.17 Kbp in total) coding for different types of bioactive compounds that hint to several possible ecological roles in the sharpnose pufferfish host.

Genome-based reclassification of Azospirillum brasilense Az39 as the type strain of Azospirillum argentinense sp. nov.

Strain Az39T of Azospirillum is a diazotrophic plant growth-promoting bacterium isolated in 1982 from the roots of wheat plants growing in Marcos Juárez, Córdoba, Argentina. It produces indole-3-acetic acid in the presence of l-tryptophan as a precursor, grows at 20-38 °C (optimal 38 °C), and the cells are curved or spiral-shaped, with diameters ranging from 0.5-0.9 to 1.8-2.2 µm. They contain C16 : 0, C18 : 0 and C18 : 1 ω7c/ω6c as the main fatty acids. Phylogenetic analysis of its 16S rRNA gene sequence confirmed that this strain belongs to the genus Azospirillum, showing a close relationship with Azospirillum baldaniorum Sp245T, Azospirillum brasilense Sp7T and Azospirillum formosense CC-Nfb-7T. Housekeeping gene analysis revealed that Az39T, together with five strains of the genus (Az19, REC3, BR 11975, MTCC4035 and MTCC4036), form a cluster apart from A. baldaniorum Sp245T, A. brasilense Sp7T and A. formosense CC-Nfb-7T. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) between Az39T and the aforementioned type strains revealed values below 96 %, the circumscription limit for the species delineation (ANI: 95.3, 94.1 and 94.0 %; dDDH: 62.9, 56.3 and 55.6 %). Furthermore, a phylogeny evaluation of the core proteome, including 809 common shared proteins, showed an independent grouping of Az39T, Az19, REC3, BR 11975, MTCC4035 and MTCC4036. The G+C content in the genomic DNA of these six strains varied from 68.3 to 68.5 %. Based on the combined phylogenetic, genomic and phenotypic characterization presented here, we consider that strain Az39T, along with strains Az19, REC3, BR 11975, MTCC4035 and MTCC4036, are members of a new Azospirillum species, for which the name Azospirillum argentinense sp. nov. is proposed. The type strain is Az39T (=LBPCV39T=BR 148428T=CCCT 22.01T).

Reduced n-3 and n-6 PUFA (DHA and AA) Concentrations in Breast Milk and Erythrocytes Phospholipids during Pregnancy and Lactation in Women with Obesity.

Obesity during pregnancy is a worrying public health problem worldwide. Maternal diet is critical for fatty acid (FA) placental transport and FA content in breast milk (BM). We evaluated FA composition in erythrocytes phospholipids (EP) and BM in pregnant women with (OBE, n = 30) and without (non-OBE, n = 31) obesity. Sixty-one healthy women were evaluated at their 20-24th gestational week and followed until 6th month of lactation. Diet was evaluated through a food frequency questionnaire. FA composition of EP and BM was assessed by gas-liquid chromatography. The OBE group showed lower diet quality, but total n-6 and n-3 polyunsaturated FA (PUFA), ALA, EPA, and DHA dietary intake was similar between groups. N-3 PUFA, ALA, DHA, and the n-6/n-3 PUFA ratio in EP were lower at the 6th lactation month in the OBE group. In BM, the arachidonic acid (AA) concentration was lower at the end of the lactation, and DHA content showed an earlier and constant decline in the OBE group compared to the non-OBE group. In conclusion, n-3 PUFA and AA and DHA levels were reduced in EP and BM in pregnant women with obesity. Strategies to increase n-3 PUFA are urgently needed during pregnancy and lactation, particularly in women with obesity.

Burkholderia perseverans sp. nov., a bacterium isolated from the Restinga ecosystem, is a producer of volatile and diffusible compounds that inhibit plant pathogens.

Gram-negative, aerobic, rod-shaped, non-spore-forming, motile bacteria, designated CBAS 719 T, CBAS 732 and CBAS 720 were isolated from leaf litter samples, collected in Espírito Santo State, Brazil, in 2008. Sequences of the 16S rRNA, gyrB, lepA and recA genes showed that these strains grouped with Burkholderia plantarii LMG 9035 T, Burkholderia gladioli LMG 2216 T and Burkholderia glumae LMG 2196 T in a clade of phytopathogenic Burkholderia species. Digital DNA-DNA hybridization experiments and ANI analyses demonstrated that strain CBAS 719 T represents a novel species in this lineage that is very closely related with B. plantarii. The genome sequence of the type strain is 7.57 Mbp and its G + C content is 69.01 mol%. The absence of growth on TSA medium supplemented with 3% (w/v) NaCl, citrate assimilation, ß-galactosidase (PNPG) activity, and of lipase C14 activity differentiated strain CBAS 719 T from B. plantarii LMG 9035 T, its nearest phylogenetic neighbor. Its predominant fatty acid components were C16:0, C18:1 ω7c, cyclo-C17:0 and summed feature 3 (C16:1 ω7c and/or C15:0 iso 2-OH). Based on these genotypic and phenotypic characteristics, the strains CBAS 719 T, CBAS 732 and CBAS 720 are classified in a novel Burkholderia species, for which the name Burkholderia perseverans sp. nov. is proposed. The type strain is CBAS 719 T (= LMG 31557 T = INN12T).

Effect of Exogenous Emulsifier (Lyso-Phospholipid) Supplementation in the Broiler Diet, on the Feed Intake and Growth Performance During Grower Phase

The objective of this experiment was to investigate the effect of supplementation of an exogenous emulsifier (lyso-phospholipid) in the diet of growing broilers on growth performance and digestibility. A total of 1224 Ross-308 ten day old broiler chicks were distributed into two experimental treatments in such a way that each treatment had twelve replicates with fifty-one birds per replicate. Two experimental diets were formulated with and without emulsifier supplementation according to the nutrition standards of Ross 308. Feed intake and body weight gain of the broilers were measured on a daily basis and feed conversion ratio was also calculated. Nutrient digestibility was determined on the 25th day of age. Analysis of variance under completely randomized design technique was used to analyze the data. Feed intake was increased (p 0.05) by supplementation of emulsifier in the broiler diet on theat 12th, 13th, 21th, 22th, 23th, 24th, and 25th days. Bodyweight gain was not affected (p>0.05) with or without emulsifier supplementation in the broiler diet during 11-25 days of life. However, feed conversion ratio was effected (p 0.05) by emulsifier supplementation and increased from days 21-25th. Nutrient digestibility (dry matter, fat, and crude protein) in the grower phase was decreased (p 0.05) by supplementation of emulsifier in the diet. It can be concluded that supplementation of an exogenous emulsifier in the diet did not show positive effect on the growth performance during the grower phase of broilers, while nutrient digestibility showed adverse effect. Emulsifier supplementation should be tested after 25 days of the life of broilers.

Effect of Exogenous Emulsifier (Lyso-Phospholipid) Supplementation in the Broiler Diet, on the Feed Intake and Growth Performance During Grower Phase

The objective of this experiment was to investigate the effect of supplementation of an exogenous emulsifier (lyso-phospholipid) in the diet of growing broilers on growth performance and digestibility. A total of 1224 Ross-308 ten day old broiler chicks were distributed into two experimental treatments in such a way that each treatment had twelve replicates with fifty-one birds per replicate. Two experimental diets were formulated with and without emulsifier supplementation according to the nutrition standards of Ross 308. Feed intake and body weight gain of the broilers were measured on a daily basis and feed conversion ratio was also calculated. Nutrient digestibility was determined on the 25th day of age. Analysis of variance under completely randomized design technique was used to analyze the data. Feed intake was increased (p 0.05) by supplementation of emulsifier in the broiler diet on theat 12th, 13th, 21th, 22th, 23th, 24th, and 25th days. Bodyweight gain was not affected (p>0.05) with or without emulsifier supplementation in the broiler diet during 11-25 days of life. However, feed conversion ratio was effected (p 0.05) by emulsifier supplementation and increased from days 21-25th. Nutrient digestibility (dry matter, fat, and crude protein) in the grower phase was decreased (p 0.05) by supplementation of emulsifier in the diet. It can be concluded that supplementation of an exogenous emulsifier in the diet did not show positive effect on the growth performance during the grower phase of broilers, while nutrient digestibility showed adverse effect. Emulsifier supplementation should be tested after 25 days of the life of broilers.(AU)

Diet, Plasma, Erythrocytes, and Spermatozoa Fatty Acid Composition Changes in Young Vegan Men.

There has been increasing interest in vegan diets, but how this dietary pattern regulates tissue fatty acids (FA), especially in men, is unclear. Our aim was to evaluate the effect of a vegan diet on plasma, erythrocyte, and spermatozoa FA composition in young men. Two groups consisting of 67 young (18-25 years old) men were studied. One group following an omnivore diet but did not consume fish, shellfish or other marine foods (control, n = 33), and another group following a vegan diet (vegan, n = 34) for at least 12 months were compared. Dietary intake was assessed via a food frequency questionnaire and a 24-h recall. FA composition was measured in plasma, erythrocyte phospholipids, and spermatozoa by gas-liquid chromatography. Compared to controls, the vegan group had higher reported intakes of carbohydrate, dietary fiber, vitamins (C, E, K, and folate), and minerals (copper, potassium) but lower intakes of cholesterol, trans FA, vitamins B6 , D, and B12 , and minerals (calcium, iron, and zinc). Vegan's reported a lower saturated FA and not arachidonic acid intake, both groups did not intake eicosapentaenoic acid and docosahexaenoic acid (DHA), but vegan's showed a higher alpha linolenic acid ALA intake. Vegans had higher plasma, erythrocyte phospholipid, and spermatozoa ALA, but lower levels of other n-3 polyunsaturated fatty acid (PUFA), especially DHA. Vegans were characterized by higher ALA, but lower levels of other n-3 PUFA, especially DHA in plasma, erythrocytes, and spermatozoids. The biological significance of these findings requires further study.

The dynamics of gene expression, lipid composition and DNA methylation reprogramming are different during in vitro maturation of pig oocytes obtained from prepubertal gilts and cycling sows.

This study aimed to characterize the gene expression, lipid composition and DNA methylation reprogramming during in vitro maturation (IVM) of pig oocytes with different developmental competencies. We used prepubertal gilts and cycling sows as a model to obtain oocytes with different levels of competency. We found that genes involved in lipid metabolism, SLC27A4, CPT2 and PLIN2, and DNA methylation, DNMT3A, TET1 and TET3, possessed altered transcript expression levels during IVM. Specifically, SLC27A4 mRNA (p = 0.05) increased in oocytes from cycling females, whereas CPT2 (p = 0.05), PLIN2 (p = 0.02) and DNMT3A (p = 0.02) increased in oocytes from prepubertal females during IVM. Additionally, TET3 mRNA increased during IVM in oocytes from prepubertal (p = 0.0005) and cycling females (p = 0.02). The TET1 transcript decreased (p = 0.05) during IVM in oocytes from cycling sows. Regarding lipid composition, mass spectrometry revealed a cluster of ions, with molecular masses higher than m/z 700, which comprises a group of complex phospholipids, was identified in all groups of oocytes, except in those from prepubertal gilts. With respect to DNA methylation reprogramming, it was noted that the less competent oocytes were not able to reprogramme the XIST gene during IVM. We conclude that the maternal mRNA store, lipid composition and epigenetic reprogramming are still being established during maturation and are related to oocyte competence. In addition, we propose that the methylation pattern of the XIST may be used as molecular marker for oocyte competence in pigs.

Phospholipid composition and resistance to vitrification of in vivo blastocyst of a Brazilian naturalized porcine breed / Composição de fosfolipídios e resistência à vitrificação de blastocistos produzidos in vivo de uma raça de suíno naturalizada brasileira

Piau porcine blastocysts were submitted to MALDI-TOF to identify the main phospholipids (PL). After that, in vivo blastocysts (D6) were vitrified (n=52), non-vitrified were used as control (n=42). After warming, blastocysts were in vitro cultured to assess re-expansion and hatching at 24 and 48 hours. Finally, at 48 hours, hatched blastocysts were submitted to RT-qPCR searching for BCL2A1, BAK, BAX and CASP3 genes. For MALDI-TOF, the ion intensity was expressed in arbitrary units. Blastocyst development was compared by Qui-square (P< 0.05). Among the most representative PL was the phosphatidylcholine [PC (32:0) + H]+; [PC (34:1) + H]+ and [PC (36:4) + H]+. Beyond the PL, MALDI revealed some triglycerides (TG), including PPL (50:2) + Na+, PPO (50:1) + Na+, PLO (52:3) + Na+ and POO (52:2) + Na. Re-expansion did not differ (P> 0.05) between fresh or vitrified blastocysts at 24 (33.3%; 32.7%) or 48 hours (2.4%; 13.5%). Hatching rates were higher (P< 0.05) for fresh compared to vitrified at 24 (66.7%; 15.4%) and 48 hours (97.6%; 36.0%). BAX was overexpressed (P< 0.05) after vitrification. In conclusion, Piau blastocysts can be cryopreserved by Cryotop. This study also demonstrated that the apoptotic pathway may be responsible for the low efficiency of porcine embryo cryopreservation.(AU)

Blastocistos de suínos foram submetidos ao MALDI-TOF para se identificarem os principais fosfolipídios (PL). Depois, parte destes embriões (D6) foram vitrificados (n=52), ou permaneceram frescos (grupo controle, n=42). Após o aquecimento, os blastocistos foram cultivados in vitro para se avaliar a reexpansão e a eclosão (BE) às 24 e 48 horas. Finalmente, às 48 horas, os BE foram submetidos ao RT-qPCR em busca dos genes BCL2A1, BAK, BAX e CASP3. No MALDI-TOF, a intensidade do íon foi expressa em unidades arbitrárias. O desenvolvimento embrionário foi comparado por qui-quadrado (P<0,05). Entre os PL mais representativos estavam as fosfatidilcolinas [PC (32: 0) + H] +; [PC (34: 1) + H] + e [PC (36: 4) + H] +. Além do PL, o MALDI revelou alguns triglicerídeos (TG), incluindo PPL (50: 2) + Na +, PPO (50: 1) + Na +, PLO (52: 3) + Na + e POO (52: 2) + Na. A reexpansão não diferiu (P>0,05) entre blastocistos frescos ou vitrificados às 24 (33,3%, 32,7%) e 48 horas (2,4%, 13,5%). As taxas de eclosão foram maiores (P<0,05) para o grupo fresco comparado ao vitrificado às 24 (66,7% x 15,4%) e 48 horas (97,6% x 36,0%). O BAX estava mais expresso (P<0,05) após a vitrificação. Concluindo, os blastocistos Piau podem ser criopreservados por Cryotop. Este estudo também demonstrou que a via apoptótica pode ser responsável pela baixa eficiência da criopreservação de embriões suínos.(AU)

Phospholipid composition and resistance to vitrification of in vivo blastocyst of a Brazilian naturalized porcine breed / Composição de fosfolipídios e resistência à vitrificação de blastocistos produzidos in vivo de uma raça de suíno naturalizada brasileira

Piau porcine blastocysts were submitted to MALDI-TOF to identify the main phospholipids (PL). After that, in vivo blastocysts (D6) were vitrified (n=52), non-vitrified were used as control (n=42). After warming, blastocysts were in vitro cultured to assess re-expansion and hatching at 24 and 48 hours. Finally, at 48 hours, hatched blastocysts were submitted to RT-qPCR searching for BCL2A1, BAK, BAX and CASP3 genes. For MALDI-TOF, the ion intensity was expressed in arbitrary units. Blastocyst development was compared by Qui-square (P< 0.05). Among the most representative PL was the phosphatidylcholine [PC (32:0) + H]+; [PC (34:1) + H]+ and [PC (36:4) + H]+. Beyond the PL, MALDI revealed some triglycerides (TG), including PPL (50:2) + Na+, PPO (50:1) + Na+, PLO (52:3) + Na+ and POO (52:2) + Na. Re-expansion did not differ (P> 0.05) between fresh or vitrified blastocysts at 24 (33.3%; 32.7%) or 48 hours (2.4%; 13.5%). Hatching rates were higher (P< 0.05) for fresh compared to vitrified at 24 (66.7%; 15.4%) and 48 hours (97.6%; 36.0%). BAX was overexpressed (P< 0.05) after vitrification. In conclusion, Piau blastocysts can be cryopreserved by Cryotop. This study also demonstrated that the apoptotic pathway may be responsible for the low efficiency of porcine embryo cryopreservation.(AU)

Blastocistos de suínos foram submetidos ao MALDI-TOF para se identificarem os principais fosfolipídios (PL). Depois, parte destes embriões (D6) foram vitrificados (n=52), ou permaneceram frescos (grupo controle, n=42). Após o aquecimento, os blastocistos foram cultivados in vitro para se avaliar a reexpansão e a eclosão (BE) às 24 e 48 horas. Finalmente, às 48 horas, os BE foram submetidos ao RT-qPCR em busca dos genes BCL2A1, BAK, BAX e CASP3. No MALDI-TOF, a intensidade do íon foi expressa em unidades arbitrárias. O desenvolvimento embrionário foi comparado por qui-quadrado (P<0,05). Entre os PL mais representativos estavam as fosfatidilcolinas [PC (32: 0) + H] +; [PC (34: 1) + H] + e [PC (36: 4) + H] +. Além do PL, o MALDI revelou alguns triglicerídeos (TG), incluindo PPL (50: 2) + Na +, PPO (50: 1) + Na +, PLO (52: 3) + Na + e POO (52: 2) + Na. A reexpansão não diferiu (P>0,05) entre blastocistos frescos ou vitrificados às 24 (33,3%, 32,7%) e 48 horas (2,4%, 13,5%). As taxas de eclosão foram maiores (P<0,05) para o grupo fresco comparado ao vitrificado às 24 (66,7% x 15,4%) e 48 horas (97,6% x 36,0%). O BAX estava mais expresso (P<0,05) após a vitrificação. Concluindo, os blastocistos Piau podem ser criopreservados por Cryotop. Este estudo também demonstrou que a via apoptótica pode ser responsável pela baixa eficiência da criopreservação de embriões suínos.(AU)

Proposal of Pedobacter nototheniae sp. nov., isolated from the spleen of a black rock cod (Notothenia coriiceps, Richardson 1844) from the Chilean Antarctica.

A pink pigmented, Gram-negative, rod-shaped, non-spore-forming bacterium (strain 36B243T), was isolated from the spleen of a black rock cod (Notothenia coriiceps, Richardson 1844) in the Chilean Antarctica. Strain 36B243T has a 5.26 Mb chromosome with a DNA G + C content of 35.4 mol%. The draft genome includes the prediction and annotation of 4585 coding genes, and 46 tRNA, 1 tmRNA, and 2735 hypothetical proteins. Phylogenetic analysis based on the 16S rRNA gene sequence placed strain 36B243T into the genus Pedobacter with high sequence similarity to the type strains of Pedobacter sandarakinus (97.5%) and Pedobacter petrophilus (97.1%). Sequence similarities to type strains of all other current Pedobacter species were below 97.1%. Predominant fatty acids are summed feature 3 (C16:1ω7c and/or C16:1ω6c) and iso-C15:0 followed by iso-C17:0 3-OH and C16:0. The major respiratory quinone was menaquinone MK-7. The polar lipid profile contained the major lipids phosphatidylethanolamine, five unidentified aminolipids, two lipids lacking a functional group and two minor glycolipids and one lipid lacking a functional group. An alkali-stable lipid was present. The polyamine pattern contained the predominant compound sym-homospermidine. Characterization by 16S rRNA gene sequence analysis, physiological parameters, pigment analysis, ubiquinone, polar lipid, and fatty acid composition revealed that strain 36B243T represents a new species of the genus Pedobacter. For this reason, we propose the name Pedobacter nototheniae sp. nov. with the type strain 36B243T (= LMG 30634T = CCM 8855T = CIP 111622T).

Buttiauxella chrysanthemi sp. nov., isolated from a chrysanthemum plantation in Brazil.

A novel Gram-negative, rod-shaped, non-motile bacterium, designated C1BT was isolated from a soil sample of a chrysanthemum plantation in Campinas, Brazil. Strain C1BT formed white colonies on BHI medium, it produces acid from D-lactose, D-mannose, D-arabinose, but does not produce from D-adonitol, m-inositol, D-melibiose, D-raffinose and D-sorbitol and it is negative for lysine and ornithine decarboxylase, phenylalanine deaminase, and citrate. Phylogenetic analysis based on 16S rRNA and rpoB genes sequences showed that strain C1BT has a similarity of 98.2 and 96.8% with different species of Buttiauxella genus. Major fatty acids were C16:0, summed features 4 (C16:1 ω7c and iso-C15:0 2OH), summed features 7 (C18:1 ω7c, C18:1 ω9t, and/or C18:1 ω12t), C17:0 cyclo, summed features 3 (iso-C16:1 I and C14:0 3OH) and C14:0. The mole percent of G+C was 49.6 mol%. Based on these results, a new species Buttiauxella chrysanthemi is proposed. The type strain is C1BT (= CPQBA 1120/15T = CMRVSP5791T).

Rhodococcus psychrotolerans sp. nov., isolated from rhizosphere of Deschampsia antarctica.

A novel actinobacterium, designated strain CMAA 1533T, was isolated from the rhizosphere of Deschampsia antarctica collected at King George Island, Antarctic Peninsula. Strain CMAA 1533T was found to grow over a wide range of temperatures (4-28 °C) and pH (4-10). Macroscopically, the colonies were observed to be circular shaped, smooth, brittle and opaque-cream on most of the culture media tested. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain CMAA 1533T belongs to the family Nocardiaceae and forms a distinct phyletic line within the genus Rhodococcus. Sequence similarity calculations indicated that the novel strain is closely related to Rhodococcus degradans CCM 4446T, Rhodococcus erythropolis NBRC 15567T and Rhodococcus triatomae DSM 44892T (≤ 96.9%). The organism was found to contain meso-diaminopimelic acid, galactose and arabinose in whole cell hydrolysates. Its predominant isoprenologue was identified as MK-8(H2) and the polar lipids as diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol and phosphatidylinositol mannosides. The major fatty acids were identified as Summed feature (C16:1 ω6c and/or C16:1 ω7c), C16:0, C18:1 ω9c and 10-methyl C18:0. The G+C content of genomic DNA was determined to be 65.5 mol%. Unlike the closely related type strains, CMAA 1533T can grow at 4 °C but not at 37 °C and was able to utilise adonitol and galactose as sole carbon sources. Based on phylogenetic, chemotaxonomic and physiological data, it is concluded that strain CMAA 1533T (= NRRL B-65465T = DSM 104532T) represents a new species of the genus Rhodococcus, for which the name Rhodococcus psychrotolerans sp. nov. is proposed.

Proton NMR characterization of intact primary and metastatic melanoma cells in 2D & 3D cultures.

OBJECTIVE: To characterize the differences between the primary and metastatic melanoma cell lines grown in 2D cultures and 3D cultures. METHODS: Primary melanoma cells (WM115) and metastatic melanoma cells (WM266) extracted from a single donor was cultured in 2D as well as 3D cultures. These cells were characterized using proton NMR spectrometry, and the qualitative chemical shifts markers were identified and discussed. RESULTS: In monolayer culture (2D), we observed one qualitative chemical shift marker for primary melanoma cells. In spheroid cultures (3D), we observed nine significant chemical shifts, of which eight markers were specific for primary melanoma spheroids, whereas the other one marker was specific to metastatic melanoma spheroids. This study suggests that the glucose accumulation and phospholipid composition vary significantly between the primary and metastatic cells lines that are obtained from a single donor and also with the cell culturing methods. 14 qualitative chemical shift markers were obtained in the comparison between monolayer culture and spheroids cultures irrespective of the differences in the cell lines. Among which 4 were unique to monolayer cultures whereas 10 chemical shifts were unique to the spheroid cultures. This study also shows that the method of cell culture would drastically affect the phospholipid composition of the cells and also depicts that the cells in spheroid culture closely resembles the cells in vivo. CONCLUSION: This study shows the high specificity of proton NMR spectrometry in characterizing cancer cell lines and also shows the variations in the glucose accumulation and phospholipid composition between the primary and metastatic melanoma cell lines from the same donor. Differences in the cell culture method does plays an important role in phospholipid composition of the cells.