Background: This study was undertaken to analyse the OA1 gene (GPR143) and its involvement in a Spanish family presenting with nystagmus, a common symptom of X‐linked ocular albinism (XLOA).
...Methods: DNA samples from the index case and eight relatives were analysed by multiplex ligation‐dependent probe amplification (MLPA). Sequence analysis and restriction assay were used to confirm the results. In addition, an analysis of a STR located in intron 1 of the OA1 gene (OA‐CA) was performed.
Results: The father of the proband presented with nystagmus, a feature consistent with XLOA. Mutation screening by multiplex ligation‐dependent probe amplification and sequence analysis of the exon 2 of the OA1 gene led to the identification of the novel p.Glu129fsX35 (g.5815delA) mutation in two affected males and four carrier females. Three relatives were found to be non‐mutated. The deletion detected resulted in a truncated protein 35 codons downstream and generated a new restriction site for the XcmI endonuclease. Additionally, microsatellite analysis showed co‐segregation with the disease in the family.
Conclusions: A novel deletion in the OA1 gene was identified in a Spanish family with ocular albinism. The mutation detected is likely a loss‐of‐function alteration. To the best of our knowledge, we describe the first Spanish family known to present with XLOA due to mutations in the OA1 gene.
Secreted Frizzled Related Proteins (SFRPs) are soluble molecules capable of modulating Wnt signalling. Different lines of evidence indicate that SFRP activity is related with the development and ...function of the retina photoreceptor cells as well as with their apoptotic degeneration associated with the onset of different cases of retinal dystrophy (RD). Because the genetic causes of many retinal dystrophies still need to be determined, we have asked whether mutations in the SFRP genes might be associated with retinal dystrophies.
Here we describe the genomic structure of SFRP1, SFRP2, and SFRP5 and a mutational screening of SFRP1 in 325 individuals affected by various non X-linked forms of inherited retinal disorders.
Three polymorphic variants were identified.
Our data, so far, exclude SFRP1 as a molecular cause of RD, since two out of three genetic variants of the gene were present in both RD patients and normal population.
Urine is a highly suitable biological matrix for metabolomics studies. Total collection for 24-h periods is the gold standard as it ensures the presence of all metabolites excreted throughout the ...day. However, in animal studies, it presents limitations related to animal welfare and also due to alterations of the metabolome originating from the use of acid for preventing microbial growth or microbial contamination. In this study, we investigated whether spot urine collection is a practical alternative to total collection for metabolomic studies in lactating cows. For this purpose, we collected urine samples from 4 lactating Holstein cows fed 4 diets in a 4 × 4 Latin square design. Urine was collected for 24 h using a collecting device (i.e., total collection) or collected once per day 4 h after the morning feeding (i.e., spot urine collection). Dietary treatments differed by the amount of nitrogen content (high vs. low) and by the nature of the energy (starch vs. fiber). Urine metabolome was analyzed by 2 untargeted complementary methods, nuclear magnetic resonance and hydrophilic-interaction liquid chromatography (HILIC) coupled to a time-of-flight mass spectrometer, and by 1 targeted method, HILIC–tandem mass spectrometry. Although sampling technique had an effect on the abundance of metabolites detected, spot urine samples were equally capable of showing differences in urine metabolome than samples from total collection. When considering nitrogen levels in the diet, the robustness and precision for discriminating high- and low-nitrogen diets was equally achieved with both sampling techniques. A total of 22 discriminant metabolites associated with the N level of diets were identified from untargeted HILIC coupled to a time-of-flight mass spectrometer (n = 9) and nuclear magnetic resonance (n = 11), and 2 from targeted HILIC–tandem mass spectrometry. Alternatively, starch or fiber in the diet induced less changes in the metabolome that were not clearly discriminated independently of the sampling technique. We concluded that spot urine collection can successfully reveal differences in the urine metabolome elicited by dietary N levels and be used as a substitute of total urinary 24-h collection for metabolomic studies.
Nitrogen use efficiency is an important index in ruminants and can be indirectly evaluated through the N isotopic discrimination between the animal and its diet (DELTA.sup.15 N.sub.animal-diet). The ...concentration and source of N may determine both the extent of the N isotopic discrimination in bacteria and N use efficiency. We hypothesised that the uptake and release of ammonia by rumen bacteria will affect the natural .sup.15 N enrichment of the bacterial biomass over their substrates (DELTA.sup.15 N.sub.bacteria-substrate) and thereby further impacting DELTA.sup.15 N.sub.animal-diet . To test this hypothesis, two independent in vitro experiments were conducted using two contrasting N sources (organic vs inorganic) at different levels either in pure rumen bacteria culture incubations (Experiment #1) or in mixed rumen cultures (Experiment #2). In Experiment #1, tryptone casein or ammonium chloride were tested at low (1 mM N) and high (11.5 mM N) concentrations on three rumen bacterial strains (Fibrobacter succinogenes, Eubacterium limosum and Xylanibacter ruminicola) incubated in triplicate in anaerobic batch monocultures during 48h. In Experiment #2 mixed rumen cultures were incubated during 120 h with peptone or ammonium chloride at five different levels of N (1.5, 3, 4.5, 6 and 12-mM). In experiment #1, DELTA.sup.15 N.sub.bacteria-substrate was lowest when the ammonia-consumer bacterium Fibrobacter succinogenes was grown on ammonium chloride, and highest when the proteolytic bacterial strain Xylanibacter ruminicola was grown on tryptone. In experiment #2, DELTA.sup.15 N.sub.bacteria-substrate was lower with inorganic (ammonium chloride) vs organic (peptone) N source. A strong negative correlation between DELTA.sup.15 N.sub.bacteria-substrate and Rikenellaceae_RC9_gut_group, a potential fibrolytic rumen bacterium, was detected. Together, our results showed that DELTA.sup.15 N.sub.bacteria-substrate may change according to the balance between synthesis of microbial protein from ammonia versus non-ammonia N sources and confirm the key role of rumen bacteria as modulators of DELTA.sup.15 N.sub.animal-diet.
Nitrogen (N) isotopic discrimination (i.e. the difference in natural .sup.15 N abundance between the animal proteins and the diet; DELTA.sup.15 N) is known to correlate with N use efficiency (NUE) ...and feed conversion efficiency (FCE) in ruminants. However, results from the literature are not always consistent across studies, likely due to isotopic discrimination pathways that may differ with the nature of diets. The objective of the present study was to assess at which level, from rumen to tissues, DELTA.sup.15 N originates and becomes related to NUE and FCE in fattening yearling bulls when they are fed two contrasted diets. Twenty-four Charolais yearling bulls were randomly divided into two groups and fed during 8 months, from weaning to slaughter, either 1) a high starch diet based on corn silage supplying a balanced N to energy ratio at the rumen level (starch) or 2) a high fiber diet based on grass silage supplying an excess of rumen degradable N (fiber). All animals were slaughtered and samples of different digestive pools (ruminal, duodenal, ileal and fecal contents), animal tissues (duodenum, liver and muscle), blood and urine were collected for each animal. Ruminal content was further used to isolate liquid-associated bacteria (LAB), protozoa and free ammonia, while plasma proteins were obtained from blood. All samples along with feed were analyzed for their N isotopic composition. For both diets, the digestive contribution (i.e. the N isotopic discrimination occurring before absorption) to the DELTA.sup.15 N observed in animal tissues accounted for 65 #177; 11%, leaving only one third to the contribution of post-absorptive metabolism. Concerning the DELTA.sup.15 N in digestive pools, the majority of these changes occurred in the rumen (av. DELTA.sup.15 N = 2.12 #177; 0.66%), with only minor .sup.15 N enrichments thereafter (av. DELTA.sup.15 N = 2.24 #177; 0.41%), highlighting the key role of the rumen on N isotopic discrimination. A strong, significant overall relationship (n = 24) between DELTA.sup.15 N and FCE or NUE was found when using any post-absorptive metabolic pool (duodenum, liver, or muscle tissues, or plasma proteins; 0.52 r 0.73; P less than or equal to 0.01), probably as these pools reflect both digestive and post-absorptive metabolic phenomena. Fiber diet compared to starch diet had a lower feed efficiency and promoted higher (P less than or equal to 0.05) DELTA.sup.15 N values across all post-absorptive metabolic pools and some digestive pools (ruminal, duodenal, and ileal contents). The within-diet relationship (n = 12) between DELTA.sup.15 N and feed efficiency was not as strong and consistent as the overall relationship, with contrasted responses between the two diets for specific pools (diet x pool interaction; P less than or equal to 0.01). Our results highlight the contrasted use of N at the rumen level between the two experimental diets and suggests the need for different equations to predict FCE or NUE from DELTA.sup.15 N according to the type of diet. In conclusion, rumen digestion and associated microbial activity can play an important role on N isotopic discrimination so rumen effect related to diet may interfere with the relationship between DELTA.sup.15 N and feed efficiency in fattening yearling bulls.
The efficiency with which ruminants convert feed to desirable products is difficult to measure under normal commercial settings. We explored the use of potential biological markers from easily ...obtainable samples, that is, blood, hair, and feces, to characterize potential causes of divergent efficiency when considered as residual feed intake (RFI) or feed conversion efficiency (FCE). A total of 54 Charolais bulls, 20 in period 1 and 34 in period 2, were examined for individual dry matter intake (DMI) and growth. Bulls were offered a diet of 70:30 wrapped grass silage to concentrate for 99 d. At the conclusion of the test period, blood samples were collected for the determination of vitamins B2 and B6, and plasma used for the determination of metabolites, natural isotopic 15N abundance (15N NIA, expressed as δ15N ‰) and fractionation (Δ15Nplasma proteins‑diet and Δ13Cplasma proteins‑diet) and near-infrared spectroscopy (NIRS). Feces were analyzed by NIRS. Bulls were slaughtered at 15–17 months of age and carcass characteristics determined. Bulls were ranked according to RFI with extremes (SD ± 0.5; n = 31) classified as either efficient (Neg-RFI) or inefficient (Pos-RFI). Extreme bulls were then classified for FCE (high vs low FCE), changing the groups. Pos-RFI bulls consumed 14% more feed than Neg-RFI bulls for the same level of weight gain. Low FCE bulls tended to eat more, but had lower weight gains than high FCE bulls. No differences were detected in carcass conformation, fat scores, hot carcass weight, or dressing percentage. Yet, heart and bladder weights were heavier in Pos-RFI, and rumen weight tended to be heavier in Pos-RFI bulls. RFI did not affect bulk 15N or 13C fractionation. A negative correlation was observed between FCE and Δ15Nplasma proteins‑diet. Inefficient bulls (Pos-RFI) had higher δ15N in glycine compared to Neg-RFI bulls. Similarly, metabolomic analysis showed a tendency for concentrations of glycine and sarcosine to be elevated in Pos-RFI bulls, whereas aspartic acid and carnosine tended to be elevated, and serine tended to be lower in High FCE. Among vitamins, only flavin adenine dinucleotide concentration was higher in the blood of bulls with High FCE. These results suggest that the two feed efficiency metrics differ in the underlying mechanisms of metabolism, where RFI is driven by differences in the energetic requirements of visceral organs and the extent of AA catabolism.
The use of biomarkers for feed conversion efficiency (FCE), such as Nitrogen isotopic discrimination (Δ
N), facilitates easier measurement and may be useful in breeding strategies. However, we need ...to better understand the relationship between FCE and Δ
N, particularly the effects of differences in the composition of liveweight gain and rumen N metabolism. Alongside measurements of FCE and Δ
N, we estimated changes in body composition and used dietary treatments with and without nitrates, and rumen metagenomics to explore these effects. Nitrate fed steers had reduced FCE and higher Δ
N in plasma compared to steers offered non-nitrate containing diets. The negative relationship between FCE and Δ
N was strengthened with the inclusion of fat depth change at the 3
lumbar vertebrae, but not with average daily gain. We identified 1,700 microbial genes with a relative abundance >0.01% of which, 26 were associated with Δ
N. These genes explained 69% of variation in Δ
N and showed clustering in two distinct functional networks. However, there was no clear relationship between their relative abundances and Δ
N, suggesting that rumen microbial genes contribute little to Δ
N. Conversely, we show that changes in the composition of gain (fat accretion) provide additional strength to the relationship between FCE and Δ
N.
The C60 polyarenes 4, 5, 18 a, and 18 b have been synthesized from truxene by triple alkylation at C5, C10, and C15 followed by a palladium‐catalyzed intramolecular arylation. The synthesis of ...“crushed fullerene” C60H30 (2) is the most efficient reported to date and proceeds in 33 % overall yield.
A triple palladium‐catalyzed arylation, which occurs with concomitant dehydrogenation, has been used to synthesize the C60H30 polyarene benzo1,2‐e:3,4‐e′:5,6‐e′′tribenzolacephenanthrylene (see scheme). Similarly, a derivative functionalized at C3, C13, and C23 has been synthesized by the same strategy.
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