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Original Research ARTICLE
Abundance and Diversity of Hydrogenotrophic Microorganisms in the Infant Gut before the Weaning Period Assessed by Denaturing Gradient Gel Electrophoresis and Quantitative PCR
- Facoltà di Scienze Agrarie, Alimentari e Ambientali, Istituto di Microbiologia, Università Cattolica del Sacro Cuore, Piacenza, Italy
Delivery mode (natural vs. cesarean) and feeding type (breast vs. formula feeding) are relevant factors for neonatal gut colonization. Biomolecular methods have shown that the ecological structure of infant microbiota is more complex than previously proposed, suggesting a relevant presence of unculturable bacteria. It has also been postulated that among unculturable bacteria, hydrogenotrophic populations might play a key role in infant health. Sulfate-reducing bacteria (SRB), acetogens, and methanogenic archaea use hydrogenotrophic pathways within the human colon. However, to date, few studies have reported detection of hydrogenotrophic microorganisms in newborns, possibly because of limitations on available group-specific, culture-independent quantification procedures. In the present work, we analyzed 16 fecal samples of healthy babies aged 1–6 months by means of quantitative PCR (qPCR) targeting the 16S rRNA or metabolic functional genes and by denaturing gradient gel electrophoresis (DGGE). qPCR data showed quantifiable levels of methanogens, SRB, and acetogens in all samples, indicating that the relative abundances of these microbial groups were not affected by delivery mode (natural vs. caesarian). DGGE revealed a high prevalence of the Blautia genus within the acetogenic bacteria despite strong interindividual variability. Our preliminary results suggest that hydrogenotrophic microorganisms, which have been a neglected group to date, should be included in future ecological and metabolic studies evaluating the infant intestinal microbiota.
Hydrogenotrophic microorganisms inhabiting the gut microbiota of humans and non-human animals are involved in a mutualistic relationship. Three major types of H2-consuming microorganisms colonize the human colon, namely, methanogenic archaea, sulfate-reducing bacteria (SRB), and acetogens (1). The common limitation in characterizing these low-abundance microbial communities is related to difficulty in their cultivation, but DNA-based techniques allow detection of microorganisms that are not easily cultured.
Microbiota development is strongly influenced by delivery mode (natural vs. cesarean) and feeding mode (breast vs. formula) (2). To date, only limited and fragmented information is available concerning the influence of delivery and feeding mode on the occurrence of hydrogenotrophic microorganisms. Breastfeeding has been positively associated with CH4 production but negatively associated with CH3SH and H2S (3). On the other hand, soy-based formula has been linked to high CH4 and H2S levels (3). In an early study, methanogens were detected in 10 out of 40 children (aged 3 months to 10 years), but their levels could not be precisely measured because the 16S rRNA gene density was below the assay's lower detection limit (4). By using quantitative PCR (qPCR), Palmer et al. (5) detected archaeal 16S rRNA gene sequences in 7 out of 14 samples from 1-year-old infants. Using a PCR assay targeting the mcrA gene, a functional gene encoding the methyl coenzyme M reductase, Mihajlovski et al. (6) reported the presence of methanogens in a newborn. Archaeal presence also has been identified in feces samples collected from birth to age 2.5 years (7). Within the archaeal group, Methanobrevibacter smithii is the dominant species detected in the human gut microbiota (8). Of interest, M. smithii was detected in all fecal specimens collected from 16 children up to the second year of life, including one sample gathered from a 2-week-old infant (9). Previous studies found no evidence of a possible correlation between intestinal archaeal colonization in newborns and dietary habits (10), but a potential, direct mother-to-child transmission has been hypothesized (5).
Concerning SRB, Fite et al. (11) reported their occurrence in 10 infants (aged 12 months) by means of qPCR targeting the 16S rRNA gene. SRB were also detected in 40 newborn stool samples collected from the neonatal period until age 24 months (12). Furthermore, the presence of SRB was reported in 15% of 12 children aged 3 months to 10 years, based on results with a real-time PCR protocol targeting the adenosine-5′-phosphosulfate reductase gene (4). More recently, De Palma et al. (13) described increased values for SRB from age 7 days to 4 months in 20 healthy babies with at least one relative who had celiac disease, as assessed by fluorescent in situ hybridization. The distribution of SRB has also been studied using primers targeting the dsrA gene, which encodes the alpha subunit of the dissimilatory sulfite reductase (DSR). The ubiquity of DSR in all known SRB microorganisms and its highly conserved nucleotide sequence make this gene suitable for evaluating the quantity and diversity of SRB in the human gut (14). In a recent paper, Pham et al. (15) investigated the occurrence of lactate-utilizing bacteria in 40 term infants, aged 2 weeks to 6 months, delivered either by vaginal birth or a C-section. In this study, SRB were enumerated by both plate count and qPCR. SRB were detected in 71% of infants at 2 weeks by means of plate count, and their levels increased with age. SRB were detected in 33, 23, 19, and 23% of babies at 2 weeks, 1 month, 3 months, and 6 months, respectively, when evaluated by qPCR.
Acetogens are obligate anaerobic bacteria that use the acetyl-CoA (Wood–Ljungdahl) pathway to synthesize acetyl-CoA from CO2 and H2 (16). Cultivation-based studies have estimated that the number of acetogens in adult human feces ranges from 102 to 108 colony-forming units/g (17). This group consists of metabolically versatile and phylogenetically diverse microorganisms. To date, more than 100 acetogenic species representing 22 genera have been isolated, most belonging to the genera Acetobacterium and Clostridium (18). In addition, some acetogenic microorganisms isolated from human feces are members of the Clostridium cluster XIVa, also known as the C. coccoides group (19), which contains a large number of butyrate-producing species (20). Moreover, some species belonging to Ruminococcus and Clostridium have been reclassified within the genus Blautia (21). The characterization of the polyphyletic distribution of acetogens and the identification at the species level using 16S rDNA-based molecular approaches is problematic because of their heterogenic composition. Recently, two functional genes—fhs, which encodes formyl tetrahydrofolate synthetase (22), and acsB, which encodes acetyl-CoA synthase (23)—have been identified as reliable molecular tools for acetogenic diversity studies. To date, acetogenic bacteria in infant fecal specimens have not been investigated.
Archaeal and bacterial diversity can be characterized using the previously described 515F/806R primer set targeting the V4 region of the 16S rRNA gene (24). However, more recent studies (25, 26) indicate that concentrations around 104 copies per microliter of this target are required to obtain reliable amplicon sequencing data for each bacterial population. In infant fecal samples, Archaeal and SRB levels are likely to fall below this cutoff and, consequently, result in no or reduced PCR product, thereby not allowing the comprehensive profiling of gut microbiota.
The aim of the present study was to assess the presence of hydrogenotrophic populations in the infant gut microbiota of 16 babies aged 1–6 months by means of qPCR and denaturing gradient gel electrophoresis (DGGE). In addition, we evaluated whether or not delivery mode influenced the dynamics of microbial colonization.
Materials and Methods
Sixteen term infants were investigated (age range: 1–6 months; mean: 2.96 months; SD: 1.35 months), and one fecal sample was collected for each subject. Babies were delivered either vaginally (n = 8) or by cesarean section (n = 8). No antibiotic treatment was provided during the 4 weeks before analysis. Samples were obtained as part of a previous study performed by Coppa et al. (27). Fecal samples were collected in 2009 at the Department of Pediatrics of the General Hospital of Ascoli Piceno and at the Neonatal Intensive Care Unit, Department of Pediatrics, of the University of Turin. The current study was conducted in compliance with the Helsinki Declaration; each mother signed an informed written consent. The Ethics Committee of the "Ospedali Riuniti" University Hospital, Polytechnic University of Marche, Ancona (Italy), reviewed and approved the study.
DNA Extraction
Stool samples were stored at −80°C until used. The samples were thawed at room temperature, and total DNA was extracted from 50 mg (wet weight) of feces using the FastDNA™ SPIN Kit for Soil (MP Biomedicals, Switzerland) according to the manufacturer's instructions (28). Genomic DNA was eluted with 100 µl of elution buffer and its quality verified by agarose gel electrophoresis.
To verify the absence of environmental contamination, a negative control reaction was included in the extraction step. Test sample was replaced by DNA-free water, which underwent the same extraction process that was used for stool specimens. DNA concentration was determined using the Qubit HS dsDNA fluorescence assay (Life Technologies, Carlsbad, CA, USA). Purified DNA was stored at −20°C until used.
Quantitative PCR
The hydrogenotrophic populations were investigated by means of qPCR with previously described primers (22, 23, 29–32), as reported in Table 1. Six different qPCR assays were performed to specifically quantify total archaea and methanogens (16S rRNA and mcrA genes, respectively), SRB (dsrA and aps genes), and acetogens (acsB and fhs genes). Additional qPCR reactions were carried out to quantify the Blautia genus and the Clostridium cluster XIVa, which also includes relevant acetogenic Blautia spp (33, 34). and total bacteria (35) (Table 1). All qPCR assays were performed in the StepOnePlus™ Real-Time PCR System (Applied Biosystems Japan, Tokyo, Japan) by using the KAPA SYBR® FAST qPCR Kit Master Mix 2X (Biolab Scientifics Instruments SA, Switzerland) or the KAPA Probe® FAST qPCR Kit Master Mix 2X (Biolab Scientifics Instruments SA, Switzerland). Tenfold serial dilutions of genomic DNA isolated from reference strains were included in each experiment to generate the standard curves; all samples including the negative control for DNA extraction and a no-template control for PCR to confirm the absence of the qPCR water contamination were processed under the conditions reported in Table 1.
Primers for DGGE analysis of the acetogens group are not available, so we used the primer pair targeting the acsB gene described by Gagen et al. (23) and modified the ACS_f primer by adding the GC-clamp. The PCR products obtained with these primers were loaded onto 8% (w/v) polyacrylamide gels (37.5/1, acrylamide/bis-acrylamide) with 20–70% linear DNA-denaturing gradients. Electrophoresis was carried out at 100 V and 60°C for 18 h in an INGENYphor 2 × 2 System (INGENYphor, Goes, Netherlands). We also used the primer pairs described by Maukonen et al. (36) to amplify the V6 region of the 16S rRNA gene of the C. coccoides–Eubacterium rectale group, a bacterial cluster that also includes some acetogenic microorganisms, among them Blautia spp. The PCR products obtained with the CcocF-GC-CcocR primers (36) were analyzed by DGGE following the methodological conditions described by Sagheddu et al. (37). Predominant bands were excised, re-amplified, sequenced (BMR Genomics, Padova, Italy), and then aligned with the GenBank database (http://www.ncbi.nlm.nih.gov/) by BLAST (38) and the blast algorithm and the Ribosomal Database Project by the Sequence Match tool (39). PCR-DGGE profiles were analyzed by Fingerprinting II SW software (Bio-Rad Laboratories, Hercules, CA, USA). Dendrograms were generated based on the Pearson's correlation coefficient by means of the Unweighted Pair Group Method with Arithmetic Mean algorithm (UPGMA).
Data normality was checked using the Shapiro–Wilk test, and homoscedasticity was assessed using Levene's test. Since qPCR data were not normally distributed, they were log-transformed prior to statistical analysis. Differences among the two tested groups were analyzed by means of t-test for independent groups (R version 3.1.2) (R Core Team, 2014) (40).
qPCR Quantification of Hydrogenotrophic Microorganisms
The cohort of 16 infants under investigation included two different experimental groups based on delivery mode. The groups were as follows: samples 1–8, C-section delivered and samples 9–16, vaginally delivered babies (Table 2).
Table 2. Quantification of hydrogenotrophic populations, Blautia spp., and Clostridium XIVa cluster by quantitative PCR.
Quantitative PCR assays were performed to evaluate the numbers of acetogens, Clostridium cluster XIVa, Blautia spp., total archaea, methanogens, SRB, and total bacteria in fecal samples. For these bacterial groups, qPCR data indicated that fecal levels were very similar across all infants (Table 2). We found a higher abundance of acetogens, as assessed both by acsB and fhs gene quantification, compared with both methanogenic archaea and SRB. In addition in our samples, the abundances of Clostridium cluster XIVa and Blautia spp. were much more homogeneous, suggesting that most of the Clostridium cluster XIVa members were represented by species within the Blautia genus. No statistically significant difference was found between experimental groups for any bacterial group tested.
PCR-DGGE Analysis of Acetogenic Bacteria in the Infant Gut
Acetogens emerged as the dominant functional bacterial group among hydrogenotrophic microorganisms in the infant gut, so we attempted to assess the composition of the acetogenic bacterial community through PCR-DGGE analysis targeting the acsB gene. This analysis produced DNA bands that could not be identified at the species level although they were all assigned to the genus Blautia (data not shown).
Although the Clostridium cluster XVIa is a phylogenetic group that includes many butyrate producers and some acetogenic microorganisms, considering the abundance of the genus Blautia in our samples, we attempted to improve the taxonomic classification of the Blautia species occurring in our stool samples by using PCR primers targeting the whole Clostridium cluster XIVa. The resulting DGGE profiles were markedly different among infants and strongly influenced by interindividual variability rather than by the delivery and feeding conditions (Figure 1). Taxonomic assignment indicated that the genus Blautia was prevalent in all fecal samples, with dominant species represented by Blautia luti, Blautia producta, Blautia wexlerae, Blautia hansenii, Lachnoanaerobaculum orale, Dorea formicigenerans, and Ruminococcus gnavus, as reported in Table 3. Of interest, in one sample, a band corresponding to Hungatella effluvii was identified (Table 3). We detected the presence of B. wexlerae in eight fecal samples (50%), B. luti in two (12.5%), B. producta in seven (43.75%), and B. hansenii in three (18.75%). The distribution of Blautia spp. was not correlated with the delivery mode, thus confirming the qPCR results.
Figure 1. (A) PCR-denaturing gradient gel electrophoresis (DGGE) profiles of 16 samples obtained using the primers CcocFGC-CcocR. Samples are subdivided by delivery mode (n = 1–8 C-section; n = 9–16 vaginal). Bands with letters were sequenced after re-amplification, and the corresponding identities were obtained by alignment in GenBank as reported in Table 3. (B) Dendrogram constructed on DGGE patterns by software analysis measured by the Pearson's correlation coefficient with the UPGMA algorithm. Columns, respectively, indicate delivery mode and infant age expressed in months.
Table 3. Identification of bacteria belonging to the Clostridium XIVa group based on denaturing gradient gel electrophoresis (DGGE) profiles (see also Figure 1A).
Hydrogenotrophic populations are present in the infant intestinal ecosystem, but their detection in infant fecal samples is hampered by their low relative abundance and by experimental limitations related to the recovery of purified high-quality DNA from infant fecal samples. Previous studies conducted on adult humans reported that 30–60% of Western people harbor methanogenic bacteria in their gut (41). More recently, some studies have described higher percentages of adult carriers (42). Methane production during childhood has been demonstrated to start around the third year of life and then to increase with age (43). Stool sample analyses, however, have revealed that methane is produced by 15.3% of 6-month-old infants at concentrations higher than 2 ppm (44). An early study estimated that approximately 108 methanogenic microorganisms per gram of dry weight of stool are needed to generate enough methane to be detectable by breath analysis (45). Additionally, direct competition among methanogens, SRB, and acetogens may occur for the common substrate H2. As reported, ~50% of healthy humans carry intestinal SRB, and several studies have described their presence at different ages in the gut lumen (4, 46) and the distal gut mucosa (14). In addition, SRB are postulated to be positively associated with inflammation (47). In contrast, few studies have assessed the presence and distribution of acetogenic bacteria in the human gastrointestinal tract.
A deep understanding of the diversity of bacteria inhabiting the infant gut microbiota is relevant for human gut ecology and future nourishment research. The major result of our study was that all infants harbored detectable levels of methanogens, acetogens, and SRB, as evaluated using specific qPCR for the 16S rRNA gene or the functional metabolic genes mcrA, acsB, fhs, dsrA, and aps. We detected about 104–105 copies/g of feces of the 16S rRNA (total archaea) and mcrA genes (methanogens), respectively, 103–104 copies/g of feces of the dsrA and aps genes (SRB), and 108–109 copies/g of feces of the acsB gene, and 109–1010 copies/g of feces of the fhs gene (acetogens), respectively. Considering the numbers of total bacteria present in our samples (about 1010–1011 16S rRNA gene copies/gram of feces), hydrogenotrophic microorganisms and in particular acetogens seem to represent a not negligible fraction of the infant gut microbiota.
The consistent recovery of hydrogenotrophic populations in all samples probably depended on the DNA extraction method performed or primer pairs used for the qPCR analyses. As Dridi et al. emphasized (9), the DNA extraction method exerts great influence on the recovery of archaea, suggesting that mechanical lysis with beads could overcome the presence of the K-resistant proteinase cell wall of methanogenic archaea. As in our study, Fite et al. (11) and Hopkins et al. (12), using a bead beating extraction protocol, recovered the presence of SRB in all fecal samples of babies tested. With a different DNA extraction system, Stewart et al. (4) reported the presence of methanogens and SRB, respectively, in 25 and 15% of fecal samples using previously described primers targeting the 16S rRNA (48, 49) and the aps (50) gene, respectively. With the same DNA extraction protocol, Palmer et al. (5) detected about 103–106 archaeal 16S rRNA gene copies/gram of wet feces in 7 out of 14 fecal samples using universal primers for the 16S rRNA gene (51, 52). These inconsistencies suggest that further efforts are needed to develop reliable molecular tools for hydrogenotrophic microorganism investigations. Notably, a recent review by Carbonero et al. (53) concluded that the occurrence of the functional genes acsB and fhs is not automatically associated to reductive acetogenesis. On the contrary, mcrA and dsrA genes seem to be reliable markers of both relative abundance and activity of methanogenic and SRB communities, respectively.
Comparison between the two DGGE analyses revealed that the 16S rRNA gene better described the complexity of the acetogenic distribution compared to the functional gene acsB. The DGGE analyses of the 16S rRNA gene allowed for identification of a greater number of acetogenic bacteria species compared to the DGGE for the acsB gene. In contrast, it was not possible to perform the DGGE analyses for the SRB and methanogens because their levels fell below the detection limit of PCR-DGGE, which ranges from 104 to 108 colony-forming units/ml (54). As a result, the presence of these microorganisms was quantifiable by qPCR, but conventional PCR did not produce any amplification fragment. In a recent review, Flint et al. (55) pointed out the necessity of investigating functional rather than phylogenetic marker genes. The study of metabolism-related genes will provide additional information allowing a comprehensive analysis of the gut microbiota. Unfortunately, in our case, only the DGGE analysis of the 16S rRNA gene gave positive results.
In this study, we failed to detect Blautia spp. 16S rRNA gene sequences by DGGE in only 1 of the 16 stool specimens, and in 2 out of 16, the band corresponding to the Blautia genus was faint; nevertheless, the identification of this genus was possible by qPCR. The Blautia genus was recognized in 93.75% of infants by DGGE and in all samples by qPCR in babies under age 6 months. Our results are in agreement with those reported by Kurakawa et al. (33), who described Blautia spp. predominance in the human intestinal Clostridium XVIa group regardless of age in three groups (32 children aged 3.2 ± 0.1 years, 32 healthy adults aged 39 ± 11 years, 32 healthy elderly aged 82 ± 6 years) (33). In recent work, Touyama et al. (56) described the presence of B. wexlerae and B. luti in the fecal samples of 12 healthy Japanese adults. The present study underlines the real problem of the possible underestimation of neglected hydrogenotrophic populations.
Another interesting finding was the recovery of a band corresponding to H. effluvii. This microorganism was recently isolated from an industrial treatment plant (57), and to the best of our knowledge, this study is the first time that it has been recovered in infant human samples. The difficulty of detecting these microorganisms is higher in infant specimens compared to adult samples, mainly because of lower levels of these microorganisms in infant specimens.
This pilot study also presents some limitations that require mention. First is the small number of recruited infants, and further studies on larger cohorts of babies are needed to define the putative role of hydrogenotrophic microorganisms in the infant gut microbiota. Since our study involved only a restricted number of samples, the major inferences that can be drawn from our results are related to the methodological approaches which allowed us to detect hydrogenotrophic populations at a quantifiable level in all infant fecal samples. The possibility of enumerating and identifying hydrogenotrophic species would be a major goal. To achieve such an aim, the punctual setting of DNA extraction protocols and the development of highly specific and efficient sets of primers with a good coverage would represent a crucial step to gain deeper insight into their actual ecological impact.
Ethics Statement
The current study was conducted in compliance with the Helsinki Declaration; each mother signed an informed written consent. The Ethics Committee of the "Ospedali Riuniti" University Hospital, Polytechnic University of Marche, Ancona (Italy), reviewed and approved the study.
Author Contributions
VS drafted the manuscript, collected samples, and performed DNA extraction and quantitative PCR. VP jointly led the study and revised the manuscript. FM performed the denaturing gradient gel electrophoresis and the cluster analysis and revised the manuscript. LM conceived and designed the study and revised the manuscript. All the authors read and approved the final manuscript.
Conflict of Interest Statement
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
This paper was a partial fulfillment of the requirements for the Ph.D. course of Valeria Sagheddu, whose Ph.D. grant has been supported by Mead Johnson Nutrition.
This work was supported by funding from Mead Johnson Nutrition (MJN) West Lloyd Expressway Evansville, IN, USA. VS has been the recipient of a doctoral research fellowship funded by MJN. The funder played no role in the preparation of the manuscript.
1. Nakamura N, Lin HC, McSweeney CS, MacKie RI, Rex Gaskins H. Mechanisms of microbial hydrogen disposal in the human colon and implications for health and disease. Annu Rev Food Sci Technol (2010) 1:363–95. doi:10.1146/annurev.food.102308.124101
2. Azad MB, Konya T, Maughan H, Guttman DS, Field CJ, Chari RS, et al. Gut microbiota of healthy Canadian infants: profiles by mode of delivery and infant diet at 4 months. CMAJ (2013) 185:385–94. doi:10.1503/cmaj.121189
3. Jiang T, Suarez FL, Levitt MD, Nelson SE, Ziegler EE. Gas production by feces of infants. J Pediatr Gastroenterol Nutr (2001) 32:534–41. doi:10.1097/00005176-200105000-00009
4. Stewart JA, Chadwick VS, Murray A. Carriage, quantification, and predominance of methanogens and sulfate-reducing bacteria in faecal samples. Lett Appl Microbiol (2006) 43:58–63. doi:10.1111/j.1472-765X.2006.01906.x
5. Palmer C, Bik EM, DiGiulio DB, Relman DA, Brown PO. Development of the human infant intestinal microbiota. PLoS Biol (2007) 5:e177. doi:10.1371/journal.pbio.0050177
6. Mihajlovski A, Doré J, Levenez F, Alric M, Brugère J-F. Molecular evaluation of the human gut methanogenic archaeal microbiota reveals an age-associated increase of the diversity. Environ Microbiol Rep (2010) 2:272–80. doi:10.1111/j.1758-2229.2009.00116.x
7. Koenig JE, Spor A, Scalfone N, Fricker AD, Stombaugh J, Knight R, et al. Succession of microbial consortia in the developing infant gut microbiome. Proc Natl Acad Sci U S A (2011) 108(Suppl):4578–85. doi:10.1073/pnas.1000081107
8. Gaci N, Borrel G, Tottey W, O'Toole PW, Brugère JF. Archaea and the human gut: new beginning of an old story. World J Gastroenterol (2014) 20:16062–78. doi:10.3748/wjg.v20.i43.16062
9. Dridi B, Henry M, El Khéchine A, Raoult D, Drancourt M. High prevalence of Methanobrevibacter smithii and Methanosphaera stadtmanae detected in the human gut using an improved DNA detection protocol. PLoS One (2009) 4:e7063. doi:10.1371/journal.pone.0007063
10. Dridi B, Raoult D, Drancourt M. Archaea as emerging organisms in complex human microbiomes. Anaerobe (2011) 17:56–63. doi:10.1016/j.anaerobe.2011.03.001
11. Fite A, Macfarlane GT, Cummings JH, Hopkins MJ, Kong SC, Furrie E, et al. Identification and quantitation of mucosal and faecal Desulfovibrios using real time polymerase chain reaction. Gut (2004) 53:523–9. doi:10.1136/gut.2003.031245
12. Hopkins MJ, Macfarlane GT, Furrie E, Fite A, Macfarlane S. Characterisation of intestinal bacteria in infant stools using real-time PCR and northern hybridisation analyses. FEMS Microbiol Ecol (2005) 54:77–85. doi:10.1016/j.femsec.2005.03.001
13. De Palma G, Capilla A, Nadal I, Nova E, Pozo T, Varea V, et al. Interplay between human leukocyte antigen genes and the microbial colonization process of the newborn intestine. Curr Issues Mol Biol (2010) 12:1–10.
14. Nava GM, Carbonero F, Croix JA, Greenberg E, Gaskins HR. Abundance and diversity of mucosa-associated hydrogenotrophic microbes in the healthy human colon. ISME J (2012) 6:57–70. doi:10.1038/ismej.2011.90
15. Pham VT, Lacroix C, Braegger CP, Chassard C. Early colonization of functional groups of microbes in the infant gut. Environ Microbiol (2016) 18:2246–58. doi:10.1111/1462-2920.13316
16. Drake HL. Acetogenesis, acetogenic bacteria, and the acetyl-CoA 'wood/ljungdahl' pathway: past and current perspectives. In: Drake HL, editor. Acetogenesis (Chapman & Hall Microbiology Series). US: Springer (1994). p. 3–60.
17. Doré J, Pochart P, Bernalier A, Goderel I, Morvan B, Rambaud JC. Enumeration of H2-utilizing methanogenic archaea, acetogenic and sulfate-reducing bacteria from human feces. FEMS Microbiol Ecol (1995) 17:279–84. doi:10.1016/0168-6496(95)00033-7
18. Drake HL, Gößner AS, Daniel SL. Old acetogens, new light. Ann N Y Acad Sci (2008) 1125:100–28. doi:10.1196/annals.1419.016
19. Bernalier A, Willems A, Leclerc M, Rochet V, Collins MD. Ruminococcus hydrogenotrophicus sp. nov., a new H2CO2-utilizing acetogenic bacterium isolated from human feces. Arch Microbiol (1996) 166:176–83. doi:10.1007/s002030050373
20. Duncan SH, Louis P, Flint HJ. Cultivable bacterial diversity from the human colon. Lett Appl Microbiol (2007) 44:343–50. doi:10.1111/j.1472-765X.2007.02129.x
21. Liu C, Finegold SM, Song Y, Lawson PA. Reclassification of Clostridium coccoides, Ruminococcus hansenii, Ruminococcus hydrogenotrophicus, Ruminococcus luti, Ruminococcus productus and Ruminococcus schinkii as Blautia coccoides gen. nov., comb. nov., Blautia hansenii comb. nov., Blautia hydroge. Int J Syst Evol Microbiol (2008) 58:1896–902. doi:10.1099/ijs.0.65208-0
22. Leaphart AB, Lovell CR. Recovery and analysis of formyltetrahydrofolate synthetase gene sequences from natural populations of acetogenic bacteria. Appl Environ Microbiol (2001) 67:1392–5. doi:10.1128/AEM.67.3.1392-1395.2001
23. Gagen EJ, Denman SE, Padmanabha J, Zadbuke S, Al Jassim R, Morrison M, et al. Functional gene analysis suggests different acetogen populations in the bovine rumen and tammar wallaby forestomach. Appl Environ Microbiol (2010) 76:7785–95. doi:10.1128/AEM.01679-10
24. Caporaso JG, Lauber CL, Walters WA, Berg-Lyons D, Huntley J, Fierer N, et al. Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms. ISME J (2012) 6:1621–4. doi:10.1038/ismej.2012.8
25. Rubin BE, Sanders JG, Hampton-Marcell J, Owens SM, Gilbert JA, Moreau CS. DNA extraction protocols cause differences in 16S rRNA amplicon sequencing efficiency but not in community profile composition or structure. Microbiologyopen (2014) 3:910–21. doi:10.1002/mbo3.216
26. Hoshino T, Inagaki F. Correction: application of stochastic labeling with random-sequence barcodes for simultaneous quantification and sequencing of environmental 16S rRNA genes. PLoS One (2017) 12:e0173546. Erratum in: PLoS One (2017) 12(1):e0169431. doi:10.1371/journal.pone.0173546
27. Coppa GV, Gabrielli O, Zampini L, Galeazzi T, Ficcadenti A, Padella L, et al. Oligosaccharides in 4 different milk groups, Bifidobacteria, and Ruminococcus obeum. J Pediatr Gastroenterol Nutr (2011) 53:80–7. doi:10.1097/MPG.0b013e3182073103
28. Jost T, Lacroix C, Braegger C, Chassard C. Stability of the maternal gut microbiota during late pregnancy and early lactation. Curr Microbiol (2014) 68:419–27. doi:10.1007/s00284-013-0491-6
29. Yu Y, Lee C, Kim J, Hwang S. Group-specific primer and probe sets to detect methanogenic communities using quantitative real-time polymerase chain reaction. Biotechnol Bioeng (2005) 89:670–9. doi:10.1002/bit.20347
30. Denman SE, Tomkins NW, McSweeney CS. Quantitation and diversity analysis of ruminal methanogenic populations in response to the antimethanogenic compound bromochloromethane. FEMS Microbiol Ecol (2007) 62:313–22. doi:10.1111/j.1574-6941.2007.00394.x
31. Christophersen CT, Morrison M, Conlon MA. Overestimation of the abundance of sulfate-reducing bacteria in human feces by quantitative PCR targeting the Desulfovibrio 16S rRNA gene. Appl Environ Microbiol (2011) 77:3544–6. doi:10.1128/AEM.02851-10
32. Leloup J, Loy A, Knab NJ, Borowski C, Wagner M, Jørgensen BB. Diversity and abundance of sulfate-reducing microorganisms in the sulfate and methane zones of a marine sediment, Black Sea. Environ Microbiol (2007) 9:131–42. doi:10.1111/j.1462-2920.2006.01122.x
33. Kurakawa T, Ogata K, Matsuda K, Tsuji H, Kubota H, Takada T, et al. Diversity of intestinal Clostridium coccoides group in the Japanese population, as demonstrated by reverse transcription-quantitative PCR. PLoS One (2015) 10:e0126226. doi:10.1371/journal.pone.0126226
34. Rinttilä T, Kassinen A, Malinen E, Krogius L, Palva A. Development of an extensive set of 16S rDNA-targeted primers for quantification of pathogenic and indigenous bacteria in faecal samples by real-time PCR. J Appl Microbiol (2004) 97:1166–77. doi:10.1111/j.1365-2672.2004.02409.x
35. Nadkarni M, Martin FE, Jacques NA, Hunter N. Determination of bacterial load by real-time PCR using a broad range (universal) probe and primer set. Microbiology (2002) 148:257–66. doi:10.1128/JCM.40.5.1698
36. Maukonen J, Mättö J, Satokari R, Söderlund H, Mattila-Sandholm T, Saarela M. PCR DGGE and RT-PCR DGGE show diversity and short-term temporal stability in the Clostridium coccoides-Eubacterium rectale group in the human intestinal microbiota. FEMS Microbiol Ecol (2006) 58:517–28. doi:10.1111/j.1574-6941.2006.00179.x
37. Sagheddu V, Patrone V, Miragoli F, Puglisi E, Morelli L. Infant early gut colonization by Lachnospiraceae: high frequency of Ruminococcus gnavus. Front Pediatr (2016) 4:57. doi:10.3389/fped.2016.00057
38. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, et al. Gapped BLAST and PS I-BLAST: a new generation of protein database search programs. Nucleic Acids Res (1997) 25:3389–402. doi:10.1093/nar/25.17.3389
39. Maidak BL, Larsen N, Mccaughey MJ, Overbeek R, Olsen GJ, Fogel K, et al. The ribosomal database project. Nucleic Acids Res (1994) 22:3485–7. doi:10.1093/nar/22.17.3485
40. R Development Core Team. R: A Language and Environment for Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing (2014). Available from: http://www.r-project.org/
41. Levitt MD, Furne JK, Kuskowski M, Ruddy J. Stability of human methanogenic flora over 35 years and a review of insights obtained from breath methane measurements. Clin Gastroenterol Hepatol (2006) 4:123–9. doi:10.1016/j.cgh.2005.11.006
42. Dridi B, Henry M, Richet H, Raoult D, Drancourt M. Age-related prevalence of Methanomassiliicoccus luminyensis in the human gut microbiome. APMIS (2012) 120:773–7. doi:10.1111/j.1600-0463.2012.02899.x
43. Sahakian AB, Jee SR, Pimentel M. Methane and the gastrointestinal tract. Dig Dis Sci (2010) 55:2135–43. doi:10.1007/s10620-009-1012-0
44. Belson A, Shetty AK, Yorgin PD, Bujanover Y, Peled Y, Dar MH, et al. Colonic hydrogen elimination and methane production in infants with and without infantile colic syndrome. Dig Dis Sci (2003) 48:1762–6. doi:10.1023/A:1025595029795
45. Weaver GA, Krause JA, Miller TL, Wolin MJ. Incidence of methanogenic bacteria in a sigmoidoscopy population: an association of methanogenic bacteria and diverticulosis. Gut (1986) 27:698–704. doi:10.1136/gut.27.6.698
46. Loubinoux J, Valente FM, Pereira IA, Costa A, Grimont PA, Le Faou AE. Reclassification of the only species of the genus Desulfomonas, Desulfomonas pigra, as Desulfovibrio piger comb. nov. Int J Syst Evol Microbiol (2002) 52:1305–8. doi:10.1099/ijs.0.02175-0
47. Devkota S, Wang Y, Musch MW, Leone V, Fehlner-Peach H, Nadimpalli A, et al. Dietary-fat-induced taurocholic acid promotes pathobiont expansion and colitis in Il10-/- mice. Nature (2012) 487:104–8. doi:10.1038/nature11225
48. Casamayor EO, Massana R, Benlloch S, Øvreås L, Díez B, Goddard VJ, et al. Changes in archaeal, bacterial and eukaryal assemblages along a salinity gradient by comparison of genetic fingerprinting methods in a multipond solar saltern. Environ Microbiol (2002) 4:338–48. doi:10.1046/j.1462-2920.2002.00297.x
49. Takai K, Horikoshi K. Rapid detection and quantification of members of the archaeal community by quantitative PCR using fluorogenic probes. Appl Environ Microbiol (2000) 66:5066–72. doi:10.1128/AEM.66.11.5066-5072.2000
50. Deplancke B, Hristova KR, Oakley HA, McCracken VJ, Aminov R, Mackie RI, et al. Molecular ecological analysis of the succession and diversity of sulfate-reducing bacteria in the mouse gastrointestinal tract. Appl Environ Microbiol (2000) 66:2166–74. doi:10.1128/AEM.66.5.2166-2174.2000
51. DeLong EF. Archaea in coastal marine environments. Proc Natl Acad Sci U S A (1992) 89:5685–9. doi:10.1073/pnas.89.12.5685
52. Lepp PW, Brinig MM, Ouverney CC, Palm K, Armitage GC, Relman DA. Methanogenic archaea and human periodontal disease. Proc Natl Acad Sci U S A (2004) 101:6176–81. doi:10.1073/pnas.0308766101
53. Carbonero F, Benefiel AC, Gaskins HR. Contributions of the microbial hydrogen economy to colonic homeostasis. Nat Rev Gastroenterol Hepatol (2012) 9:504–18. doi:10.1038/nrgastro.2012.85
54. Temmerman R, Scheirlinck I, Huys G, Swings J. Culture-independent analysis of probiotic products by denaturing gradient gel electrophoresis. Appl Environ Microbiol (2003) 69:220–6. doi:10.1128/AEM.69.1.220-226.2003
55. Flint HJ, Karen P, Louis P, Duncan SH. The role of the gut microbiota in nutrition and health the role of the gut microbiota in nutrition and health. Nat Rev Gastroenterol Hepatol (2012) 9:577–89. doi:10.1038/nrgastro.2012.156
56. Touyama M, Jin JS, Kibe R, Hayashi H, Benno Y. Quantification of Blautia wexlerae and Blautia luti in human faeces by real-time PCR using specific primers. Benef Microbes (2015) 6:583–90. doi:10.3920/BM2014.0133
57. Kaur S, Yawar M, Kumar PA, Suresh K. Hungatella effluvii gen. nov., sp. nov., an obligately anaerobic bacterium isolated from an effluent treatment plant, and reclassification of Clostridium hathewayi as Hungatella hathewayi gen. nov., comb. nov. Int J Syst Evol Microbiol (2014) 64:710–8. doi:10.1099/ijs.0.056986-0
Keywords: hydrogenotrophs, babies, gut microbiota, Blautia, quantitative PCR
Citation: Sagheddu V, Patrone V, Miragoli F and Morelli L (2017) Abundance and Diversity of Hydrogenotrophic Microorganisms in the Infant Gut before the Weaning Period Assessed by Denaturing Gradient Gel Electrophoresis and Quantitative PCR. Front. Nutr. 4:29. doi: 10.3389/fnut.2017.00029
Received: 15 December 2016; Accepted: 09 June 2017;
Published: 26 June 2017
Edited by:Aldo Corsetti, University of Teramo, Italy
Reviewed by:Franck Carbonero, University of Arkansas, United States
Benoit Chassaing, Georgia State University, United States
Copyright: © 2017 Sagheddu, Patrone, Miragoli and Morelli. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Vania Patrone, firstname.lastname@example.com


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HARNESS Racing New South Wales (HRNSW) Stewards conducted an inquiry on Thursday 9 August 2018 into licensed driver Mr Leigh Sutton recording an alcohol concentration in excess of 0.02% on a breath analyser at the time he presented to drive at the Bathurst harness meeting on Sunday 29 July 2018. Mr Sutton pleaded guilty to a charge pursuant to Australian Harness Racing Rule (AHRR) 250(1)(a) as follows: AHHR 250. (1) A driver commits an offence if:- (a) A sample taken from him is found upon analysis to contain a substance banned by Rule 251, AHHR 251. The following substances and/or their metabolites, artifacts, and isomers are declared as banned substances in drivers when present in a urine sample (unless otherwise stated) at a concentration above the applicable cut-off level:- (c) All diuretics (0μg/L): Probenecid: (0μg/L): Alcohol (at a concentration in excess of 0.02% on a breath analyser): Mr Sutton's driver's licence was wholly suspended for a period of six months to commence from 29 July 2018, the date upon which he was stood down. Mr Sutton was also provided with an opportunity to have the suspension reduced to five months if he completed sessions with an alcohol counsellor approved by HRNSW. Mr Sutton was informed that an application would be considered by HRNSW Stewards for him to return to trackwork two weeks prior to the end of the relevant suspension period. In considering penalty Stewards were mindful of the following: Mr Sutton's first offence for such matter; Seriousness of the offence; The concentration of alcohol recorded being 0.095%; Mr Sutton's guilty plea, licence history, personal and financial subjective facts. Harness Racing NSW (HRNSW) is the controlling body for harness racing in New South Wales with responsibility for commercial and regulatory management of the industry including 33 racing clubs across the State. HRNSW is headed by a Board of Directors and is independent of Government. To arrange an interview or for further information please contact: MICHAEL PRENTICE | INTEGRITY MANAGER (02) 9722 6600 • firstname.lastname@example.com GRANT ADAMS | CHAIRMAN OF STEWARDS (02) 9722 6600 • firstname.lastname@example.com
They've come from all over for a swing at Sunday's Seelite Windows and Doors Victoria Trotters Derby, determined to salute in front of a boisterous crowd. New Zealand trainer Cran Dalgety's trotter Chevron Express and fellow Kiwi President Roydon, who is now in Kerryn Manning's stable, will bring some international flair when they have an almighty crack at the $75,000 feature. Chevron Express is among a fearsome group of fillies who have amassed for the three-year-old feature, who equate for seven of the 13 Derby entrants and include Anton Golino's Cardigan-based trio Dance Craze, Une Belle Allure and Nieta. For Manning, Sunday presents a chance to capture one of the few pieces of silverware that has thus far escaped her grasp. President Roydon has spent six weeks with team Manning, having previously raced with Mark Purdon's all conquering All Stars team over the Tasman, where only highly-rated Enghien prevented him breaking through at Group 1 level. The Great Western trainer-driver held the reins as the gelding made his Australian debut at Tabcorp Park Melton on Friday night, when President Roydon led and did the bulk of the work only to be picked off by Garshaway in the dying stages. "He's a funny horse," Manning said of President Roydon, who has drawn gate two for Sunday's Group 1. "He's not the most wonderful to drive, he's a bit green and awkward at times, but he tries really hard." Manning said he should improve from his Australian debut, when he "over raced a little bit in front and was found wanting a little in the last 50". "He hasn't had a start for a month and the run would have done him good and he seems to have pulled up well. The distance (2690m) will suit, he's a strong horse." The "good little sit-sprint horse" that whooshed past her was improving Garshaway, Myola-based trainer Graeme McDermott's gelding who has drawn gate 13. He will be driven by New South Wales' Leigh Sutton, who took the steer after Manning was unavailable due to her commitment with President Roydon and saluted at Melton at $19.60 on the TAB. "I come down with my trotter (last Friday) so they offered me the drive and said if he goes all right I can drive him in the Derby," Sutton said. "It will be nice to have a drive in a Group 1 race." The Victoria Trotters Derby takes place at 3.54pm. Michael Howard (HRV Media/Communications Co-Ordinator)
Reinsman Nathan Jack will honour the late Tiffany Murray by wearing her driving pants at Echuca Harness Racing Club's annual Pacing Cup meeting on Saturday night. Tiffany, a popular local harness racing figure, died in 2011 after a brave battle with cancer. Jack knew Tiffany well through his good mate and Tiffany's partner, Leigh Sutton. Sutton is also a harness racing driver and he and Tiffany have a daughter, Milla. Jack will also donate $100 per winning drive to the Cure Brain Cancer Foundation – and $200 should he win the Echuca Cup – and Sutton will match it dollar for dollar. It means the organisation set up by Tiffany's neurosurgeon, Charlie Teo, will benefit $200 per winner and $400 if Jack's horse, Smudge Bromac, can win the main event. "I knew her quite well. I was good mates with Leigh," Jack said. "It's definitely a cause close to my heart." The club will also run the Tiffany Murray Memorial Pace (Race 4 at 8.03pm), with only female drivers invited to contest the event. The association between Jack and Smudge Bromac will hold him in good stead aboard the David Aiken trained entire on Saturday night. The nine-year-old pacer is in a rich vein of form, having recently contested the $400,000 Hunter Cup, finishing fifth beaten only 8.3 metres by Arden Rooney. "He's a great little horse, you'd love to own him," Jack said of the 31-time winner. "He's in great form. His Yarra Glen Cup win was terrific, in the Pure Steel he sat outside Uncle Wingnut and beat him by over a metre, and in the Shepparton Cup he turned for home level with For A Reason and beat him a length. "He'll find it a bit hard off the 20-metre (handicap) on the tight track but he's racing really well." The huge 10-race program at Echuca kicks off at 6.20pm with the cup to be run at 9.33pm. Loads of giveaways and entertainment will be provided, with one lucky patron winning $1000 thanks to the TAB simply by uploading their Trot Selfie to facebook.com/trotselfies. The Moama Bowling Club Echuca Pacing Cup meeting is a leg of the TAB Trots Country Cups Carnival. by Cody Winnell Related Links Fields for Echuca, Saturday 21 March 2015 Form guide for Echuca, Saturday 21 March 2015
More than 130 guests are set to enjoy a tremendous evening's entertainment as local harness racing stars are recognised at Wagga's awards dinner on Saturday night. Incorporating the Riverina titles for the Trainer, Driver, Concession Driver, Horse and Juvenile of the Year, the function is always an enjoyable affair. Champion mare Frith and New South Wales Oaks winner, Sheas A Runa, will be the recipients of the top Riverina horse awards, with Major Jules claiming the Wagga Pacer of the Year crown. Talented reinsman Leigh Sutton captured the Riverina and Wagga driving premierships, while Shaun Snudden (Riverina) and Kim Hillier (Wagga) annexed the other awards. Wagga chief executive Graeme White also revealed 65 sponsors have been placed in a draw for naming rights of this season's feature races. "Last season first-time sponsor, Ausmet Roofing, won naming rights for the Wagga Cup and there are seven new sponsors for this season," White said. "There are other sponsors who have been with the club for many years, so who wins is all a matter of having some luck on the night." Wagga's racing dates for 2014/15 include the Carnival Of Cups in March, plus several Tuesday night meetings during the warmer months, which will be something different for the region. Headlining the entertainment is Australia's leading female ventriloquist, Lindi Jane, who is bound to keep the laughs flowing on a frequent basis. Harness Racing NSW chairman Rex Horne and Wagga Mayor, Rod Kendall, will be two of the special guests. HRNSW Media
Tuesday's Bendigo harness racing fixture featured two events for the squaregaiters, with Apollos Gift and Sonofanearl winning their respective races.
Maryborough - the harness racing home of "The Redwood" played host to heats of the Crazed Australasian Breeders Crown Series on Monday July 30, with two heats for Two Year Olds and three for Three Year Olds over 2100 metres.
Victoria's biggest provincial harness racing carnival - Mildura Cup week commenced on Tuesday April 10, featuring two heats of the Park Douglas Printing Mildura Pacing Cup, supplemented by several other heats of races catering for all classes, including the Mark Gurry & Associates Cup for C4 to C8 class over 2190 metres and the Tankard Dental Mildura Guineas for Three Year Olds over the same journey.
Calder Park (Geelong) principal Ric Burchell a lover of the squaregaiter, has enjoyed plenty of happy harness racing moments over the years with his trotters of note such as Calder Sensation, Calder Strike, Calder Image and Calder Reign to name just a few, but may have something special in Metropolitan/Like A Calder colt Blitzthemcalder who was successful at start number two in taking out The Athletes Foot 2-Y-0 Trotters Handicap over 2200 metres at Bray Raceway Ballarat on Thursday March 29.
Harness racing trainers, drivers and horses from the greater Bendigo area dominated the Kilmore harness fixture on Wednesday January 4, winning half the program, including a double to the all conquering Douglas stable.
A terrific harness racing meeting was held at Tabcorp Park melton on Friday, with the most elated person on course being Pyramid Hill (Kerang) owner/breeder Teddy Carmody, following the victory of six year old Conch Deville/Boot Scootin Glory gelding Unbundled in the Tabcorp Park Melton Trotters Mobile for TMO class over 2240 metres.
Less than four days before staging their own Tiffany Murray Trust Dinner at Tabcorp Park, Harness Racing Victoria would like to acknowledge the efforts of Lyle Massey and Geoff Lucas in raising more than $42,500 for the courageous cancer sufferer in Mildura last Friday night.
Local harness racing owners Vince and Maria Sanzaro were "over the moon" after winning the $25,000 SEW-Eurodrive Geelong Rocket at Beckley Park Geelong on Saturday with former Kiwi 7-Y-0 gelding Lets Slash An Burn.
This Saturday night (September 17) the Albion Park Harness Racing Club will conduct an eight event card at headquarters in Brisbane. The night's racing will kick off with an M1 event at 6.10. Gatton based trainer Shane Sanderson has a terrific strike rate with his team of pacers.
The Nyah harness racing club held an interesting eight event card on Thursday July 28, with the honours for the night going to Echuca trainer Faye McEwan and Bendigo based reinsman Leigh Sutton who combined to land a stable treble.

subset: commoncrawl


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Out-Law News
UK Pensions Regulator publishes DB superfund 'gateway principles' guidance
Out-Law Analysis
Compulsory independent scrutiny of pre-pack sales to connected parties
Out-Law Analysis | 26 Aug 2015 | 2:56 pm | 4 min. read
There are considerable differences between the new regime and the old voluntary report, and trustees risk fines of between £500 and £2,000 if they do not comply. Trustees should make sure that they are well-placed to complete the chair's statement in advance.
The new governance standards
The new governance standards are set out in the Charges and Governance Regulations, and explained by the Pensions Regulator in new guidance. They require scheme trustees to demonstrate sufficient knowledge and understanding to effectively run the pension scheme; to ensure that financial transactions are processed promptly and accurately; and to regularly review costs and charges borne by members to ensure that these are value for money, amongst other things.
The regulations also introduced the requirement that trustees appoint a chair, which should have been done by 5 July 2015. A statement setting out how the new standards have been met, signed off by the chair, will from now on have to be included in a mandatory scheme annual report.
DC trustees will already be aware that they have had the opportunity to produce a voluntary statement along these lines by completing a standard form put together by the Pensions Regulator. This form allows you to report on the conclusions of your self-assessment of the scheme, relative to the standards described in Code of Practice 13 (59-page / 442KB PDF) and the associated guidance (33-page / 98KB PDF). The Pensions Regulator has suggested including this voluntary report with the new mandatory report and accounts, along with further details of your assessment of the scheme, if helpful.
The chair's statement must be included in the scheme's published annual report and accounts. The trustees as a group will also need to declare that they have produced the chair's statement as part of the scheme return. To emphasise the importance of this, you could be fined between £500 and £2,000 if you do not comply with the requirements.
Preparing for the chair's statement
The first chair's statement will generally be due within seven months of the end of the scheme year, unless the period that would be covered by that statement is three months or less. Once you have established when the first statement is due, it is a good idea for the trustees as a group to complete a draft statement well in advance. This will allow you to identify, and to rectify, any potential issues at an early stage.
Among the questions you should be asking yourselves before completing a draft statement are:
Your answers to some of these questions may require more detailed consideration.
Default arrangements
Even if your scheme does not describe a particular fund or strategy as its "default", it may be caught by the new governance requirements for default arrangements. These requirements do not necessarily only apply in circumstances where members have not expressed a choice as to where their contributions are allocated, as they can also apply where 80% of the contributing members or more are in the same arrangement. This might mean there is more than one default arrangement in a given scheme, and could even mean that further default arrangements emerge over time.
The strict letter of the legislation can create some surprising results. Guidance by the Department for Work and Pensions (DWP) (31-page / 531KB PDF) is there to help, but it is by no means comprehensive. It is therefore very much worth doing some detailed analysis, at least for the purpose of your first statement.
The charge cap
The 0.75% charge cap introduced by the regulations covers all administration charges and must be calculated in a particular way. The calculation may need to be adapted to make sure that any leavers in the scheme year are not charged in excess of the cap. Please also note that the percentage cap could be as low as 0.4% if a flat rate fee, or some other combination of fee, is also charged. This is also addressed in the DWP guidance. Again, some detailed analysis may well be required to ensure that you can discharge your duties.
The trustees as a group will also be required to confirm that they are meeting the charge controls as part of the scheme return, following changes by the Pensions Regulator.
Value for money
The Pensions Regulator has published material to help with the value for money assessment, but this remains a very difficult area for trustees. As with all trustee decision-making, the important thing is to follow the proper processes under trust law and to make sure the right considerations are taken into account.
The existing Code of Practice
Code of Practice 13 remains a very important guide as to how trustees should go about discharging their legal duties under pensions legislation and trust law. The new governance requirements and chair's statement build on, rather than replace, these legal duties. You should continue to make every effort to comply with Code of Practice 13 in addition to satisfying the new requirements.
A couple of areas included in the code of practice remain somewhat neglected by trustees, and the new regime provides a timely opportunity to address these. Both issues relate to existing legal requirements.
Under section 249A of the 2004 Pensions Act, trustees are required to set up and operate "adequate internal controls" to ensure the scheme is administered in accordance with the law and scheme rules. The Pensions Regulator requires you to enter into "clear and comprehensive contracts" with suppliers including clear service levels, standards of care and terms relating to data protection, data security, delegation, notice periods and termination costs and assistance in order to demonstrate compliance with this provision.
Regulation 4(7) of the 2005 Occupational Pension Schemes (Investment) Regulations requires trustees to give "due consideration" to asset protection, and to understand what would happen in the event of a problem. In particular, you are expected to establish whether the funds that they invest in qualify for the Financial Services Compensation Scheme (FSCS), and to communicate the position to members.
Understanding and managing the contract risk associated with third party relationships is an important part of complying with Code of Practice 13, and your associated legal duties to members.
Tom Barton is a pensions law expert at Pinsent Masons, the law firm behind Out-Law.com.
Out-Law News
UK Pensions Regulator publishes DB superfund 'gateway principles' guidance
Out-Law Analysis
Compulsory independent scrutiny of pre-pack sales to connected parties


text:
Telemedicine is now at the forefront of a transformation affecting doctors and the patients they serve. It has the power to affect care at every level, from pre-treatment consults to remote monitoring and postoperative treatment.
What's driving this shift is not only the exorbitant and rising costs of providing care but also access to that care is growing more problematic. It's common today to wait a week or longer to see a primary care physician; for some specialties, the wait is longer, particularly in rural communities. A recent study showed that patients travel an average of 34-minutes to reach their on-site medical appointment. Each visit tacks on another 11-minutes on average of waiting room time. The time spent traveling and waiting for treatment is a significant drain on patients, to the estimated tune of $89 billion in lost productivity and time off work. If these visits are for routine postoperative care, they may only spend a few minutes with a provider face-to-face.
Healthcare Dive says the dilemma of traveling and waiting, along with the time it takes to even get the appointment, hasn't improved in more than a decade. Ironically, the solution to these problems is as close as our smartphones. Let's examine how telemedicine can positively affect postoperative care and reduce travel and wait times along with the associated financial costs.
Telemedicine vs. Traditional Visit – Outcomes the Same for Postoperative Care
Last year, a study in the journal mHealth sought to examine the specific impact of telemedicine on postoperative treatment. While numerous clinical studies continue to be published on how the virtual visit can positively impact surgical care, routine clinical visits, and even proactive population health initiatives, this article sought to document the value of telehealth in postoperative care. The researchers noted that telemedicine in postoperative care, "demonstrated excellent clinical outcomes, enhanced patient satisfaction, increased accessibility along with reduced wait times, and cost savings for patients and health care systems."
In general, telemedicine continues to promote positive outcomes in several areas, including:
- Improved access to care.
- Reduced resource inefficiencies.
- Decreases costs.
Today, more than 50% of U.S. acute care hospitals use telemedicine and the industry is expected to exceed $30 billion in revenues this year alone. The application of telehealth for postoperative care shows that clinical outcomes are at least as high as the traditional clinical follow-up. The mHealth study pointed out:
In elective low-risk procedures like laparoscopic and open inguinal hernia repair, laparoscopic cholecystectomy, parathyroidectomy, arthroplasty and pediatric urological procedures, complication rates generally range from 0 to 4.8% which are not statistically different from the complication rates following traditional clinical follow-ups postoperatively.
Too, non-elective procedures, such as those caused by orthopedic trauma have higher rates of complication. But a 2015 study showed that there was very little difference in patient outcomes when the postoperative visits were conducted via telemedicine for postoperative orthopedic care. A systematic review of the literature stated, "No studies have reported statistically higher complication rates with telemedicine visits when compared to complication rates in traditional clinical follow-ups."
How Can Telemedicine Be Applied in Postoperative Care?
For more than a decade, telemedicine has been used in three primary ways to provide postoperative care to surgical patients:
- Scheduled follow-up visits.
Click here to read the study.
- Routine monitoring.
Click here to read the study.
- Management of postoperative issues as they arise.
Click here to read the study.
Recently, the application of telehealth for postoperative care as a replacement of traditional clinic visits has grown more popular. Automated messaging, online video conferencing, or a smartphone call is all growing in popularity.
One pilot study from 2017 tracked clinical outcomes when post-surgical wounds were monitored via a smartphone for appendectomy patients. One week after surgery, patients sent an image of their surgical wound to their clinician and completed a questionnaire. The wound histology was then analyzed by two independent physicians. On the eighth day, the patient attended a traditional visit with another physician and completed a satisfaction questionnaire.
The study found that the uses of telemedicine for postoperative wound care showed a sensitivity of 100% and a specificity of 91.6%. Further, there was agreement between the two types of evaluations (virtual and on-site) and 94% of the patients were pleased with the telemedicine visit. The study concluded:
The telemedicine-based follow-up, has proven to be feasible and safe for the evaluation of early postoperative complications. Patients reported high levels of satisfaction with the procedure. Telemedicine-based follow-up could become standard practice with the development of a specific mobile application.
Using telemedicine in postoperative treatment has also yielded effective results for:
- Text messaging of drain outputs for post-surgical breast reconstruction patients.
- Video conferencing of ileostomy outputs for post-surgical cancer patients.
- Automated system alerts to monitor post-surgical cancer patients that underwent a thoracotomy.
- A smartphone app to track medication adherence and blood pressure in post-surgical kidney transplant patients.
What's startling to note is that each of these studies, despite the variances in patient treatment, disease modalities, or even the type of telehealth application used all showed positive outcomes. Using telehealth to monitor postoperative surgical patients reduced appointment no-shows, providing patients with improved and increased monitoring of their conditions. These studies showed that patient outcomes improved in every instance of telehealth use.
Patient Satisfaction and Telehealth Use
Patient satisfaction is increasingly an imperative for all types of care, and certainly for surgical and postoperative treatment. Telehealth, as a new tool in many settings, should certainly rank highly from the patient's perspective. But patient's report they are highly satisfied with remote postoperative treatment via telehealth. This satisfaction stretches across the fields of transplant, vascular, and general surgery. Whether the clinician leveraged telehealth applications for monitoring wounds, providing education and support, tracking vital signs, or medication adherence, the mHealth study noted consistently high patient satisfaction scores.
Further, the physicians themselves report a high level of satisfaction during the use of these tools. For example:
- 88% of urologists report being satisfied with video visits for radical prostatectomy.
Click here to read the study.
- Providers report a 90% satisfaction rating for using telemedicine for postoperative burn patients.
Click here to read the study.
It's clear that telemedicine for postoperative treatment over the traditional office visit holds real potential to reshape provider workflows. Contact OrthoLive to find out how telemedicine could be used in your surgical orthopedic practice.


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Information Technology Manager
Location: Finance & Administration
Pay rate: Hiring Salary range: $59,082.23 to $76,806.91
Closing Date: Open Until Filled
This full-time, exempt position will be responsible for the management, effectiveness and impact of technology at the Durham Housing Authority. In addition to driving the operations and impact of the function, this position will also have hands-on responsibilities. The manager will ensure security of data, network access and backup systems. Plan and manage annual budgets, the efforts and services of software providers, as well as applications and infrastructure projects. Will research and implement emerging technologies pertaining to agency goals and manage the agency's technology assets. Will work closely with senior management to determine future needs, plan projects that help to achieve goals and define success metrics. Acquire and maintain an understanding of operations within the affordable housing industry. The position requires a BS in Computer Science, MIS or similar field, plus 5 years working experience as an IT Manager or related experience.


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|Review: Stielas Storhett - Skd
Label: De Profundis
Year released: 2010
Duration: 15:27
Tracks: 2
Genre: Black Metal
Rating: 4.25/5
Review online: April 18, 2010
Reviewed by: Michel Renaud
Rated 3/5 (60%) (1 Vote)
Stielas Storhett's Skd may be short in length, but it packs a lot in those 15 minutes. This is slow-paced, atmospheric, dark and depressing Black Metal that makes you feel likes it's dragging you down to hell along with it. The vocals sound like raw, raspy/screechy laments and convey an inescapable feeling of hopelessness – that's some sick, tormented stuff right there and it doesn't let go. As if that wasn't enough, the music reinforces these dark emotions. Slow-paced melancholic passages are king here and create a very depressing atmosphere that just pin you to the ground with no chance of escape. Things do get heavier and faster at times, but still as crushingly fucked up as the slower stuff. This MCD really feels much longer than 15 minutes (in a good way – that is if you can call inducing depression a good thing ;)) If you think you've been feeling too happy lately, a few repeated listens of Skd are guaranteed to make you run to the doctor for a prescription of anti-depressant medication.
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A transportation company operator whose broken-down stretch limo was involved in the horrific weekend crash upstate that killed 20 people was arrested Wednesday and charged with criminally negligent homicide.
Nauman Hussain, 28, was pulled over during a morning traffic stop and arrested in connection with the Saturday accident by state police who said he was directly responsible for the tragedy. He was a passenger in a car driven by his older brother Shahyer. Prosecutors said there were several bags in the vehicle packed with his belongings.
Officials said the cause of the crash that killed the driver, 17 passengers and two people in a packed Schoharie parking lot Saturday is still under investigation. But they said this much is clear: Hussain put a driver without the proper paperwork behind the wheel of a shoddy SUV that had no business being on the road.
"The driver he hired should not have been operating the type of vehicle involved in Saturday's crash," said State Police Superintendent George Beach. "He was cited with the violations. He knew this. The sole responsibility for that motor vehicle being on the road on Saturday rests with Nauman Hussain. That vehicle was placed out of service by the Department of Transportation in September of this year and should not have been on the road."
Sources said the vehicle was affixed with a large sticker after the failed inspection deeming it out of service. "This vehicle is NOT to be operated until repaired," the sticker read.
Hussain, 28, whose father owns the Wilton, N.Y.-based Prestige Limousine company, was pulled over along Interstate 787 in Watervliet as he was riding with his brother. It was not the first time the siblings were in a car together during a traffic stop.
About four years ago, the brothers, who look alike, pretended to be each other when they were pulled over without identification with Shahyer behind the wheel, officials said.
The two were arrested after an investigation revealed that Shahyer's license was revoked and had 28 suspensions, he'd used his brother's identity before and that combined, the duo had been ticketed more than 70 times.
Hussain's father, Shahed Hussain, is currently out of the country. His company owned the 2001 Ford Excursion that blew a stop sign, cut through the parking lot of the Apple Barrel Country Store and slammed into an embankment, killing all aboard and two pedestrians in the lot.
The company's safety record and the vehicle's history report have come under intense scrutiny since the crash. The limo failed two inspections this year, including one in March that revealed the hydraulic brakes were not working properly, officials said.
Hussain's lawyer, Lee Kindlon, told the Daily News the arrest had "jumped the gun." He said the road was more of a factor in the crash.
"It was a known problem to the state of New York," Kindlon said. "Frankly, the Department of Transportation and the state of New York are doing a great job of saying, 'Look over there, it's not our fault!' Is this day a distracter for what the state of New York knew and when they knew it? That's always going to be up for answers."
A state Transportation Department official pushed back against the claim that the intersection sparked the crash.
"This location was fully reconstructed about a decade ago and there have been only four reported accidents since reconstruction," DOT spokesman Joe Morrissey said. "In addition, we have imposed restrictions on trucks since 2013 and fully prohibited trucks in 2015."
The driver's wife, Kim Lisinicchia, told CBS that she overheard her husband, Scott, complain several times about the vehicle's safety. "I'm not going to drive this, like this," she said, quoting her husband. "You need to get me another car."
Hussain pleaded not guilty at his arraignment Wednesday night in Cobleskill Town Court and was released on $150,000 bail. Schoharie DA Susan Mallery had asked for $50,000 cash or $150,000 bond, a "higher bail and bond than usual," because "he may have dual citizenship in Pakistan and the United States," she said.
"We ask that he surrender his United States passport and Pakistani passport to the DA," Mallery added. "Our concern is that he was leaving the jurisdiction today with packed belongings."
His lawyer Kindlon said that despite numerous death threats in the past few days, Hussain was a U.S. citizen, doesn't have a Pakistani passport and "wasn't going anywhere."
Hussain, who wore a black t-shirt, gray basketball shorts and a Yankees hat at his arraignment, looked nervous and kept looking back at his brother Shahyer, a girlfriend and a friend who were sitting in the audience.
He's due back in court on Oct 23.


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Tuesday, February 27, 2018
PPG Paints Arena
TV: TVAS, SN, MSG+, AT&T SportsNet
Radio: TOJN, 105.9 'The X'
After two days off the Penguins (36-23-4) return to action tonight against the New Jersey Devils (32-22-8). In those two days off, we have seen the Flyers jump into first place in the M...
Saturday, February 24, 2018
BB&T Center
TV: FS-F, AT&T SportsNet
Radio: WQAM, 105.9 'The X'
This entrance by the nWo in the late 1990s during the Monday Night War is exactly how I feel about the Penguins anytime they enter trade talks.
As seen on Twitter yesterday, and this graphic, the Pengui...
Friday, February 23, 2018
PNC Arena
TV: TVAS, FS-CR, AT&T SportsNet
Radio: 99.9 FM 'The Fan', 105.9 'The X'
21 games remaining. The Penguins have shot up the standings, but are still in a fight and can't afford to lose games, including games against teams like the Hurricanes. Pittsburgh (35-...
Tuesday, February 20, 2018
As the trade deadline draws closer and closer, now with it being less than a week away, its time to start talking about the ideal additions to this years Penguins squad. If you've been living under a rock or have given up on the team much like superstar fan from Twitte...
Sunday, February 18, 2018
Nationwide Arena
TV: FS-O, AT&T SportsNet
Radio: 97.1FM, 105.9 'The X'
The red hot Penguins (34-22-4) will look for their fifth straight win tonight when they travel to Columbus to take on the Columbus Blue Jackets (29-25-4) tonight with puck drop set for just afte...
Saturday, February 17, 2018
PPG Paints Arena
TV: NHL Network - US, SN, AT&T Sportsnet
Radio: TSN / SportsNet,105.9 'The X'
Tonight the Penguins (33-22-4) welcome the Toronto Maple Leafs (35-19-5) into town. Right now both teams are coming into this affair on hot streaks. The Leafs have won fi...
Thursday, February 15, 2018
PPG Paints Arena
TV: TVAS, SN1, SNP, FS-W, AT&T SportsNet
Radio: KABC 790, 105.9 'The X'
Whatsup dudes and dudettes? Tonight is 90's Night at PPG Paints Arena and the piece of shit Los Angeles Kings are in town. The Kings are coming off of a drubbing by the Carolin...
Tuesday, February 13, 2018
PPG Paints Arena
TV: RDS, TSN5, AT&T SportsNet
Radio: TSN1200, 105.9 'The X'
Nine months ago these two teams met in the Eastern Conference Finals. Fast forward to the present and these two teams couldn't have gone on more different paths. Since a slow start to th...
Sunday, February 11, 2018
Scottrade Center
Radio: KMOX 1120, 105.9 'The X'
Following a 4-3 shootout loss in Dallas Friday night, the Penguins (30-22-4) will look to build a new winning streak when they take on the St. Louis Blues (34-20-3) today with puck drop set for just af...
Friday, February 9, 2018
American Airlines Center
TV: SN1, FS-SW, AT&T SportsNet
Radio: KTCK, 105.9 'The X'
The Flightless Birds of the Steel City waddle down to Dallas to take on the Stars, The Guins are coming off a 5-4 win at PPG Paints Arena against the Vegas Golden Knights and a bunch...
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Meditation is training your body in awareness while getting your mind to open up to a new, better perspective and to observe your thoughts without judgement to understand them better. It helps you in reducing stress, controlling anxiety, beating insomnia as well as making you aware of your emotional and physical health. People start to take meditation as hobby or part of their routine when hear about its benefits but may leave it in between.
This may be because of many reasons which could be distraction, lots of worries and thoughts going on in your mind, not getting time for it or being busy. These things may make you think that it's not your thing, or you are still good enough without it which makes it even difficult to do. Meditation is a very Simple Phenomenon as described by OSHO, Indian spiritual Guru.
OSHO : Meditation is a very Simple Phenomenon
The famous and admired spiritual leader, Rajnish or Osho explains meditation as a simple phenomenon which makes it even easier to understand. Osho describes, meditation Is a very simple phenomenon, when you are not doing anything at all bodily, physically, mentally, on no level, when all activity has ceased and you simply are, just being, that's what meditation is.
You don't have to practice it, just try to understand it. You can't do it, you can't rehearse it, and you have just to get it. It cannot be taught it has to be caught. And whenever you can find time for just being and drop all doing, thinking is also doing, mind, concentration is also doing, and contemplation is also doing. Whenever for any single moment, if you are not doing anything and you are just calm at your center, feeling utterly relaxed, that is meditation.
And once you have got this practice into your habit, you can remain in that state as long as you want. You can spend all 24 hours in this state of relaxation or meditation. Once you have caught this technique of being relaxed and undisturbed then slowly you can proceed in your all activities in relaxation. That is the second very important phase of meditation. When you have turned out to be mindful of the way, of how a being can stay undisturbed, at that point gradually you can begin getting things done – keeping ready that being isn't blended. Also Check Out: Exploring Life-Spiritual
In the first place, figuring out how just to be and afterward adapting little activities: cleaning the floor, scrubbing down, yet keeping yourself focused. At that point, you can start doing little complicated things. For instance, OSHO says, "I am addressing you yet my meditation not disturbed. I can continue talking, yet at my inside, there isn't even a swell. It is simply quiet, absolutely quiet".
So meditation isn't against the activity, it isn't that you need to escape from life. It just shows you another lifestyle, where you become the focal point of a violent wind. Your life goes on, it goes on extremely more strongly – with more bliss, with greater lucidity, more vision, and greater innovativeness – yet you are unapproachable, only a watcher on the slopes essentially observing all that is going on around you. Also Read: How to be Pleasant Everytime!!!
OSHO says, "You are not the practitioner, you are the watcher. That is the entire mystery of meditation that you become the watcher". Doing proceeds without anyone else level, there is no issue… slashing wood, drawing water from the well. You can do all little and enormous things. Just a single thing isn't permitted and that is, your focusing ought not to be lost. That mindfulness, that watchfulness ought to remain completely unclouded, undisturbed. Meditation is a very simple and straightforward phenomenon.


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Previously known as Park 511 and Erie Park, the A. Montgomery Ward Park is a furrific spot to spend a beautiful afternoon with your four-legged best friend. Located in the vibrant River North neighborhood, this 3-acre green space on the Chicago River draws residents with its river promenade, children's playground, and dog-friendly area.
You and Fido will find the dog-friendly area in the north section of the park, which is where the canine members of the community hang out. This section includes both on-leash and off-leash areas, giving River North's doggos ample space to stretch their legs and get some exercise. The A. Dog Park, also referred to as the Larrabee Dog Park, is a fenced-in facility where your furry pal can romp and socialize without the restriction of a leash. It features water fountains, some trees, seating for humans, and a pea gravel surface that's gentler on your pooch's paws. During the summer months, hoses are available to help pups cool off. Oh, and you get pawsome views of the river and downtown architecture, too!
Note that there is no separate enclosure for smaller canines, and doggie waste bags don't seem to be provided, so do bring your own. Leave the treats at home, though, as no food is allowed inside the dog park. As with any off-leash area in the city, your pup will need a permit and tag from a participating veterinarian before they can use the A. Montgomery Ward Dog Park. If your pooch is not in the mood to mingle, you can always take them for a stroll around the surrounding on-leash area. Just be sure to stay in the north section of Ward Park, as the south section is off-limits to doggos.
For a barking good time in River North, bring your pup to the A. Montgomery Ward Park!


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Heli-Hike the Berg Lake Trail
14.3 miles -2610 ft gain - Point-to-Point Trail
Added by Jeremy Meek
Pristine glacial lakes. Tallest mountain in the Canadian Rockies. Rope Bridges. Passing through the Valley of a Thousand Falls. Plus, all down hill? Count me in!
If you do not have the time (or energy) to backpack into Berg Lake and camp overnight, one solution is to fly into Berg Lake via Robson Helimagic's chopper and hike out. Going with this heli-hiking route allows you to experience this iconic trail without the hassle of camping and getting proper permits. If you are interested in the backpacking and camping experience, check out Morgan Lee Alain's post here.
The helicopter will drop you off near the Robson Pass Campground, which is immediately adjacent the border between Alberta and British Columbia (Jasper National Park and Mt. Robson Provincial Park). From here, you get to hike downhill 2,610' over the course of 14.3 miles. You will get to hike along the Robson River, surrounded by Hargreaves, Robson, Berg, and Mist Glaciers initially. As you get farther down the trail, you will pass by the monstrous Emperor Falls (I have witnessed backpackers taking a "shower" in the mist of Emperor Falls). Continuing on, you will pass through a variety of waterfalls and lakes as you make your way through the Valley of a Thousand Falls. Kinney Lake and the neighboring campground will be your last major point of interest along your hike out to the trailhead. Mount Robson towers over you throughout the hike; fitting considering it is the tallest mountain in the Canadian Rockies at 12,972'.
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Kevin AbernethyExplorer
🥇Top Contributor
about 3 years ago
Best Hike Ever
This trail was phenomenal! The helicopter ride started things off perfectly for us, we were able to see everything from a birds eye. You are dropped off at Robson Pass, but we took the short detour to Mum Basin to get a different view of Berg Lake. The trail was in excellent condition, and there was something to see around every corner. Permits are hard to come by if you plan on camping, but I'd recommend taking your time out here. If you are taking the helicopter ride you can make this a day full day hike, we stopped and camped at Kinney Lake just so we could really take our time around the trail. One of the best parts is the trail wasn't overly crowded, Mt Robson is just west of Jasper and it didn't have any crowds in comparison to Banff, Yoho, or Jasper. Don't skip out on this if you are in the area.
Nearby Adventures
Hike to Overlander Falls
0.6 miles 150 ft gain
Hike to Snowbird Pass, Mount Robson Provincial Park
14 miles 2300 ft gain
Backpack the Berg Lake Trail
26.1 miles 2500 ft gain
Backpack to Berg Lake
24 miles 2591.9 ft gain


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Stef Renée: Ghana Odyssey
Written by LiRon Anderson-Bell on January 27, 2017
WURD family, our own Stef Renée has spent the last week in Ghana as part of a tour arranged by the Israeli Consulate General, who invited members of the media to document the work they're doing there. Many don't know that Ghana and Israel have worked on projects together for decades; in fact, Ghana has an embassy in Ramat Gan, Israel, and Israel has an embassy in Ghana.
Stef will bring everyone up to speed on her amazing adventure during next week's MOJO, but in the meantime, please enjoy her recent photos and vlog of the journey below:
I learned my lesson about Facebook Live on my last trip. Only trying that when I know I've got excellent signal. Otherwise, you get the slightly delayed video updates. Let's roll to West Africa!
Posted by Stephanie Renée on Saturday, January 21, 2017
Getting ready for ?
Posted by Stephanie Renée on Saturday, January 21, 2017
Akwaaba means welcome! And so do complimentary slippers that will definitely be used and brought back home. #StefExodus (Kempinski Hotel Gold Coast City, Accra, Ghana)
Good morning from Accra. A quick preview of today's activities.
Posted by Stephanie Renée on Monday, January 23, 2017
A preview of the day, as we ship out for my next Ghanaian adventures.
Posted by Stephanie Renée on Monday, January 23, 2017
Royalty. Paying respects to the recently deceased Queen Mother of Ashantiland. #nofilterneeded #StefExodus (Manhyia Palace, Accra, Ghana)
The time has arrived. My first taste of official Jollof rice in Ghana. The smack talking between us Nigerians and the Ghanaians thus far has been legendary.
Last full day in Ghana. More nonprofits and NGOs to visit.
Posted by Stephanie Renée on Wednesday, January 25, 2017
Most of our media delegates, along with the Israeli Ambassador to Ghana, and the children of BASICS International. Heart is so full! #StefExodus (Chorkor Lantemaamli, Accra, Ghana)
This is it, folks. One final day of running around, building & learning, then on a ✈ back to the States late tonight. Hope to see you soon.
Posted by Stephanie Renée on Thursday, January 26, 2017
Part of the media delegation featured as guests on Metro TV's "Good Morning Ghana" today, sharing info on our journey.


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The ULTIMATE investment buys: Ten designer pieces that will look great now (and could make you rich in the future!)
- High-end clothing and accessories can make for excellent investments
- Some must-have items have seen their value double over the last ten years
- Here FEMAIL reveals the ten money-making designer pieces to snap up today
Property, stocks and gold have long been earmarked as good investments but an even better money-maker could be tucked away at the back of your wardrobe.
Clothing and accessories from the world's top designers are more than just beautiful must-have items, they also accrue value over time.
An iconic Hermès Birkin bag – as seen on the arm of Victoria Beckham and Heidi Klum, among others – bought for £3,750 in 2007 would be worth nearly twice as much if sold second-hand today.
Meanwhile a Chanel Medium Classic Flap Bag has seen its value skyrocket nearly 72 per cent between 2010 and 2015, research by Baghunter revealed.
And while only the well-heeled might already own one of these timeless pieces, Claudia Valentin, Designer Goods Expert at Prestige Pawnbrokers, has picked out 10 others that can still be snapped up – either in store or on pre-loved websites.
'While it's hard to say for definite what these items will be worth in the future – any investment comes with risk – you can look at their previous price jumps and see that they are relatively safe bets,' she said.
Here FEMAIL reveals ten investment pieces to buy now – if you have the cash.
Chanel Jumbo Classic Timeless
Retail price in 2007 £1,800; second-hand price now £3,500
Classic: Fashion designer Nicole Richie, left, and actress Jessica Alba, right, are among the celebrities who have been snapped with a Chanel Timeless bag, which comes in several sizes
Name any celebrity and chances are there's a photo of them online carrying a Chanel bag with pride – that golden chain strap has long been synonymous with the great and good.
Particular fans of the Timeless bag include Nicole Richie and Jessica Alba.
Classic: Fashion designer Nicole Richie, left, and actress Jessica Alba, right, are among the celebrities who have been snapped with a Chanel Timeless bag, which comes in several sizes
'It's called Timeless for a reason,' says Claudia. 'There have only been slight changes to the clasp since this was very first designed – it still has the same chain and buttery-soft leather that it always has.'
Rolex Submariner
Retail price in 2007 £2,000; second-hand price now £5,000
When it comes to Hollywood, Rolex has quite the pedigree.
Steve McQueen was rarely pictured without his trusty Submariner, and it's the exact model worn by Sean Connery in Dr No – if it's good enough for Bond, it's good enough for us.
'With Rolex – as with Cartier, Chanel and other big brands – you're buying the name as much as anything,' says Claudia.
'The name stands for quality, and that's why they hold their value so well.'
Iconic: David Beckham is among the stars who has been spotted with a Rolex watch
Cartier Love Bangle
Retail price in 2007 £3,500; second-hand price now £5,000
First seen at the tail end of the swinging sixties, Love Bangles show no sign of diminishing in popularity, and have graced the wrists of everyone from Pippa Middleton to Lindsay Lohan.
Claudia said: 'Cartier pieces are heirlooms, designed to hand down through the generations. The brand is so well known and trusted that even second-hand pieces are worth a lot.'
Heirlooms: Pippa Middleton, left, and Katie Holmes, right, are both fans of the Love Bangle
Hermès Birkin Handbag
Retail price in 2007 £3,750; second-hand price now £7,500
Famously created by Jane Birkin herself, after a chance meeting with Jean-Louis Dumas, who had recently taken over his family's leather goods company, when she complained aloud that bags didn't have enough pockets.
Handbag history was made, when the two designed the Birkin bag – now more famous than the actress herself.
'Apart from the fact that this is a piece that goes with any outfit, it's also wonderfully practical thanks to its numerous pockets,' says Claudia.
Must-have: Models Heidi Klum, left, and Miranda Kerr, right, show off the Hermes Birkin bag
Hermès Kelly Handbag
Retail price in 2007 £3,750; second-hand price now £7,500
Exclusive: Victoria Beckham proved her style credentials with the Hermès Kelly in 2009 +13
Exclusive: Victoria Beckham proved her style credentials with the Hermès Kelly in 2009
It's not hard to imagine why this bag has been so popular, thanks to its connections with style icon Grace Kelly, who loved the design so much she ordered six in different colours.
Thanks to her, it was often photographed in glossy magazines, clasped in her perfectly manicured hands.
'This bag is special in part because of its exclusivity,' says Claudia. 'It can only be purchased in person from an Hermès boutique.'
Louis Vuitton Mirage Speedy 30 (limited edition)
Retail price in 2007 £400; second-hand price now £1,200
We've lost count of the number of celebrities touting these monogrammed bags – from Rihanna to Kim K, SJP to Lady Gaga, everyone wants to be seen bustling through the airport with this on their arm.
'It's a timeless style that's not designed to be seasonal,' says Claudia, 'so, like all of these items, it won't date.'
Exclusive: Victoria Beckham proved her style credentials with the Hermès Kelly in 2009
Chanel women's Tweed Jacket
Retail price in 2007 £1,000; second-hand price now £2,250
Versatile: Chanel has recreated its distinctive tweed jacket in a number of styles and colours over the decades. Pictured, Poppy Delevingne, left, and Kristin Scott Thomas in two designs
Loved by Miranda Kerr, Clemence Poesy, Kate Upton – and basically every model ever – this little boxy classic smacks of Audrey Hepburn glamour.
'It's all about power dressing,' says Claudia, 'This tweed jacket has been seen on powerful women, from Jackie O to Margaret Thatcher, so wearing it tells a story about how you see yourself.'
Gucci women's 'Peyton' Embellished Riding Boots
Retail price in 2007 £400; second-hand price now £1,750
Edgy: Rita Ora in the short version of the Gucci Peyton boots, on sale now
Gucci loves its royal connections almost as much as its horsey ones – previously signing Charlotte Casiraghi, the granddaughter of Princess Grace of Monaco and superb horse rider, as a model – and its riding boots smack of Princess Anne and Zara Phillips at the polo.
They even sponsor their own riding event – the Gucci Masters – to attract the elite to their brand.
'The subtle interlocked G's on these boots are a way for the wealthy to show off their status without being garish, and they look good with most outfits,' says Claudia.
Christian Dior Haute Couture Cocktail Dress
Retail price in 2007 £1,500; second-hand price now £3,000
Any haute couture cocktail dress from Christian Dior might as well be laced with gold – they're worth lots to the right buyer thanks to their glamorous connections with red carpet big-hitters, from Jennifer Lawrence to Amal Clooney, and even Princess Diana wasn't immune to their charms.
'With haute couture you're paying for the time and dedication of the craftsmanship,' says Claudia, 'many man-hours will have gone into each individual item, and you can see the quality instantly.'
Tiffany Solitaire Diamond Ring
Retail price in 2007 £8,000; second-hand price now £10,000
Timeless: Jennifer Lawrence, left in 2013, and Amal Clooney, right in 2015, are both fans of Dior. The French designer is known for creating elegant gowns that make perfect investments
Apart from the obvious Audrey Hepburn connection, Tiffany & Co has always had star power. Richard Burton bought Elizabeth Taylor a diamond and sapphire clip, while Marilyn Monroe name-checked the brand in Diamonds Are A Girl's Best Friend.
'Such a well-known, iconic brand is very unlikely to dip in value,' says Claudia. 'It's still just as popular with celebrities today as it was back in the 1950s and 60s.'
Here, two experts offer advice to anyone thinking about curating an investment wardrobe.
Claudia Valentin said: 'The main thing to remember is to keep items in tip-top condition, and keep any dust bags, boxes and paperwork safe, as they drop in value without these.'
Brand and Style Expert Nick Ede explained that jewellery and accessories are particularly good at holding their value as 'one size fits all'.
He said: 'While dresses, coats and shoes may be fabulous and are still collectors' items, they may also be bought by people who want to wear them, so you're more limited in your willing buyers than with a size-free accessory.'
But Nick also suggested shoppers interested in investing take a look at fast fashion, pointing to designer-high street tie-ins.
He said: 'Items from H&M with Balmain were being sold for 10 times their RRP just days after they went on sale, so you need to consider whether your investment piece is a long-term classic or an instant must-have item.'
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