Longitudinal Reduction in Diversity of Maternal Gut Microbiota During Pregnancy Is Observed in Multiple Low-Resource Settings: Results From the Women First Trial.

Objective: To characterize the changes in gut microbiota during pregnancy and determine the effects of nutritional intervention on gut microbiota in women from sub-Saharan Africa (the Democratic Republic of the Congo, DRC), South Asia (India and Pakistan), and Central America (Guatemala). Methods: Pregnant women in the Women First (WF) Preconception Maternal Nutrition Trial were included in this analysis. Participants were randomized to receive a lipid-based micronutrient supplement either ≥3 months before pregnancy (Arm 1); started the same intervention late in the first trimester (Arm 2); or received no nutrition supplements besides those self-administered or prescribed through local health services (Arm 3). Stool and blood samples were collected during the first and third trimesters. Findings presented here include fecal 16S rRNA gene-based profiling and systemic and intestinal inflammatory biomarkers, including alpha (1)-acid glycoprotein (AGP), C-reactive protein (CRP), fecal myeloperoxidase (MPO), and calprotectin. Results: Stool samples were collected from 640 women (DRC, n = 157; India, n = 102; Guatemala, n = 276; and Pakistan, n = 105). Gut microbial community structure did not differ by intervention arm but changed significantly during pregnancy. Richness, a measure of alpha-diversity, decreased over pregnancy. Community composition (beta-diversity) also showed a significant change from first to third trimester in all four sites. Of the top 10 most abundant genera, unclassified Lachnospiraceae significantly decreased in Guatemala and unclassified Ruminococcaceae significantly decreased in Guatemala and DRC. The change in the overall community structure at the genus level was associated with a decrease in the abundances of certain genera with low heterogeneity among the four sites. Intervention arms were not significantly associated with inflammatory biomarkers at 12 or 34 weeks. AGP significantly decreased from 12 to 34 weeks of pregnancy, whereas CRP, MPO, and calprotectin did not significantly change over time. None of these biomarkers were significantly associated with the gut microbiota diversity. Conclusion: The longitudinal reduction of individual genera (both commensals and potential pathogens) and alpha-diversity among all sites were consistent and suggested that the effect of pregnancy on the maternal microbiota overrides other influencing factors, such as nutrition intervention, geographical location, diet, race, and other demographical variables.

Birth length is the strongest predictor of linear growth status and stunting in the first 2 years of life after a preconception maternal nutrition intervention: the children of the Women First trial.

BACKGROUND: The multicountry Women First trial demonstrated that nutritional supplementation initiated prior to conception (arm 1) or early pregnancy (arm 2) and continued until delivery resulted in significantly greater length at birth and 6 mo compared with infants in the control arm (arm 3). OBJECTIVES: We evaluated intervention effects on infants' longitudinal growth trajectory from birth through 24 mo and identified predictors of length status and stunting at 24 mo. METHODS: Infants' anthropometry was obtained at 6, 12, 18, and 24 mo after the Women First trial (registered at clinicaltrials.gov as NCT01883193), which was conducted in low-resource settings: Democratic Republic of Congo, Guatemala, India, and Pakistan. Longitudinal models evaluated intervention effects on infants' growth trajectory from birth to 24 mo, with additional modeling used to identify adjusted predictors for growth trajectories and outcomes at 24 mo. RESULTS: Data for 2337 (95% of original live births) infants were evaluated. At 24 mo, stunting rates were 62.8%, 64.8%, and 66.3% for arms 1, 2, and 3, respectively (NS). For the length-for-age z-score (LAZ) trajectory, treatment arm was a significant predictor, with adjusted mean differences of 0.19 SD (95% CI: 0.08, 0.30; P < 0.001) and 0.17 SD (95% CI: 0.07, 0.27; P < 0.001) for arms 1 and 2, respectively. The strongest predictors of LAZ at 24 mo were birth LAZ <-2 and <-1 to ≥-2, with adjusted mean differences of -0.76 SD (95% CI: -0.93, -0.58; P < 0.001) and -0.47 SD (95% CI: -0.56, -0.38; P < 0.001), respectively. For infants with ultrasound-determined gestational age (n = 1329), the strongest predictors of stunting were birth LAZ <-2 and <-1 to ≥- 2: adjusted relative risk of 1.62 (95% CI: 1.39, 1.88; P < 0.001) and 1.46 (95% CI: 1.31, 1.62; P < 0.001), respectively. CONCLUSIONS: Substantial improvements in postnatal growth are likely to depend on improved intrauterine growth, especially during early pregnancy.

Gestational weight gain in 4 low- and middle-income countries and associations with birth outcomes: a secondary analysis of the Women First Trial.

BACKGROUND: Adequate gestational weight gain (GWG) is essential for healthy fetal growth. However, in low- and middle-income countries, where malnutrition is prevalent, little information is available about GWG and how it might be modified by nutritional status and interventions. OBJECTIVE: We describe GWG and its associations with fetal growth and birth outcomes. We also examined the extent to which prepregnancy BMI, and preconception and early weight gain modify GWG, and its effects on fetal growth. METHODS: This was a secondary analysis of the Women First Trial, including 2331 women within the Democratic Republic of Congo (DRC), Guatemala, India, and Pakistan, evaluating weight gain from enrollment to ∼12 weeks of gestation and GWG velocity (kg/wk) between ∼12 and 32 weeks of gestation. Adequacy of GWG velocity was compared with 2009 Institute of Medicine recommendations, according to maternal BMI. Early weight gain (EWG), GWG velocity, and adequacy of GWG were related to birth outcomes using linear and Poisson models. RESULTS: GWG velocity (mean ± SD) varied by site: 0.22 ± 0.15 kg/wk in DRC, 0.30 ± 0.23 in Pakistan, 0.31 ± 0.14 in Guatemala, and 0.39 ± 0.13 in India, (P <0.0001). An increase of 0.1 kg/wk in maternal GWG was associated with a 0.13 cm (95% CI: 0.07, 0.18, P <0.001) increase in birth length and a 0.032 kg (0.022, 0.042, P <0.001) increase in birth weight. Compared to women with inadequate GWG, women who had adequate GWG delivered newborns with a higher mean length and weight: 47.98 ± 2.04 cm compared with 47.40 ± 2.17 cm (P <0.001) and 2.864 ± 0.425 kg compared with 2.764 ± 0.418 kg (P <0.001). Baseline BMI, EWG, and GWG were all associated with birth length and weight. CONCLUSIONS: These results underscore the importance of adequate maternal nutrition both before and during pregnancy as a potentially modifiable factor to improve fetal growth.

An atypical presentation of Salmonella enterica ser Dublin in an immunocompromised patient.

Mycotic aneurysms of the carotid artery are a rare entity that can be fatal if not diagnosed promptly. We present a 60-year-old man with a tender left-sided neck mass due to a ruptured aneurysm of the left internal carotid artery. Cultures taken intraoperatively grew Salmonella enterica ser Dublin.

Growth from Birth Through Six Months for Infants of Mothers in the "Women First" Preconception Maternal Nutrition Trial.

OBJECTIVE: To evaluate whether the fetal linear growth effects of maternal nutrition supplementation would be maintained through 6 months postnatal age. STUDY DESIGN: The Women First trial was a multicountry, individually randomized clinical trial that compared the impact of maternal nutrition supplementation initiated preconception (Arm 1) vs at ∼11 weeks of gestation (Arm 2), vs no supplement (Arm 3); the intervention was discontinued at delivery. Trial sites were in Democratic Republic of Congo, Guatemala, India, and Pakistan. Analysis includes 2421 infants born to 2408 randomized women. Primary outcome was the trajectory of length-for-age z scores (LAZ) by arm, based on assessments at birth and 1, 3, and 6 months. We fitted longitudinal models on growth from birth to 6 months using generalized estimating equations; maternal intervention effects were evaluated, adjusting for site and baseline maternal covariates. RESULTS: Linear growth for Arms 1 and 2 was statistically greater than for Arm 3 in 3 of the 4 countries, with average pairwise mean differences in LAZ of 0.25 (95% CI 0.15-0.35; P < .001) and 0.19 (95% CI 0.09-0.28; P < .001), respectively. Compared with Arm 3, average overall adjusted relative risks (95% CI) for stunting (LAZ <-2) were lower for Arms 1 and 2: 0.76 (0.66-0.87; P < .001) and 0.77 (0.67-0.88; P < .001), respectively. CONCLUSIONS: Improved linear growth in early infancy observed for the 2 intervention arms supports the critical importance of maternal nutrition before conception and in the early phase of gestation. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01883193.

Florid hypercementosis synchronous with periodontitis: a case report.

Excessive cementum formation, referred to as hypercementosis (HC), is an uncommon nonneoplastic process that principally occurs with permanent teeth. Widespread tooth involvement has been confined mostly to Paget disease of bone. Only a limited number of reports of HC coincident with periodontitis has appeared in the literature. The aim of this article is to present the clinical, radiographic, and histopathologic findings of a 44-year-old female with moderate to severe periodontitis synchronous with 22 HC-affected teeth. A list of other etiologies associated with HC is provided.

Crisflash: open-source software to generate CRISPR guide RNAs against genomes annotated with individual variation.

SUMMARY: CRISPR/Cas9 system requires short guide RNAs (sgRNAs) to direct genome modification. Most currently available tools for sgRNA design operate only with standard reference genomes, and are best suited for small-scale projects. To address these limitations, we developed Crisflash, a software tool for fast sgRNA design and potential off-target discovery, built for performance and flexibility. Crisflash can rapidly design CRISPR guides against any sequenced genome or genome sequences, and can optimize guide accuracy by incorporating user-supplied variant data. Crisflash is over an order of magnitude faster than comparable tools, even using a single CPU core, and efficiently and robustly scores the potential off-targeting of all possible candidate CRISPR guide oligonucleotides. AVAILABILITY AND IMPLEMENTATION: https://github.com/crisflash. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Ethnic and language influence on parents' perception of paediatric behaviour management techniques.

BACKGROUND: Parental preference for various behaviour management techniques (BMTs) used in paediatric dentistry has been shown to be influenced by many factors, including ethnicity. AIM: To measure parental acceptability of BMTs used in paediatric dentistry and how it is influenced by ethnicity and language. DESIGN: Parents of patients presenting to a paediatric dentistry residency clinic in Houston, Texas, USA or Medellín, Colombia watched ten video BMT vignettes and rated their acceptance on a visual analog scale (VAS). Participants were categorized into six groups based on language, ethnicity, and country of residence. RESULTS: Parental acceptance of BMTs was affected by language, ethnicity, and country of residence (P = 2.2 × 10-16 ). Ethnic groups in the USA had a mean overall acceptance rate of all BMTs. Colombians rated all BMTs less acceptable than the US cohorts (P < 0.05), with the exception of voice control, which Colombians rate less acceptable than English-speaking Caucasians and Spanish-speaking Hispanics in the USA (P < 0.05). The Colombian population were not accepting of conscious sedation, nitrous oxide, general anaesthesia, and protective stabilization. CONCLUSIONS: Parents from different ethnic groups express different preferences in BMTs. Parents continue to prefer noninvasive techniques over pharmacologic and advanced techniques, with the exception of voice control.

Gloss and Surface Roughness of Anterior Pediatric Zirconia Crowns.

PURPOSE: The purpose of this study was to determine gloss and surface roughness (Ra) of pediatric anterior zirconia crowns. METHODS: Gloss of labial and lingual surfaces of pediatric anterior zirconia crowns from three manufacturers was measured on 20 specimens using a small area gloss meter on each. Ra (µm) was measured using a contact-type surface profilometer. Data were evaluated by analysis of variance and pair-wise comparison at the 0.05 level of significance. RESULTS: There were statistically significant interactions between surface location and crown type for both gloss and Ra scores. NuSmile had higher mean gloss scores and lower mean Ra scores than both Kinder Krowns and EZCrowns. Kinder Krowns showed lower mean gloss scores and higher Ra scores than other crown groups. CONCLUSION: Among all crowns, there was a trend of higher mean gloss paired with lower mean surface roughness, and lower mean gloss paired with higher mean Ra. Hand smoothed followed by mechanically polished zirconia crowns (NuSmile) displayed the highest mean gloss and lowest mean Ra compared to hybrid polishedglazed zirconia crowns (Kinder Krowns, EZCrowns). Of the hybrid polished-glazed zirconia crowns, Kinder Krowns displayed the lowest mean gloss and highest mean Ra.

Identification of DNA hypermethylation of SOX9 in association with bladder cancer progression using CpG microarrays.

CpG island arrays represent a high-throughput epigenomic discovery platform to identify global disease-specific promoter hypermethylation candidates along bladder cancer progression. DNA obtained from 10 pairs of invasive bladder tumours were profiled vs their respective normal urothelium using differential methylation hybridisation on custom-made CpG arrays (n=12 288 clones). Promoter hypermethylation of 84 clones was simultaneously shown in at least 70% of the tumours. SOX9 was selected for further validation by bisulphite genomic sequencing and methylation-specific polymerase chain reaction in bladder cancer cells (n=11) and primary bladder tumours (n=101). Hypermethylation was observed in bladder cancer cells and associated with lack of gene expression, being restored in vitro by a demethylating agent. In primary bladder tumours, SOX9 hypermethylation was present in 56.4% of the cases. Moreover, SOX9 hypermethylation was significantly associated with tumour grade and overall survival. Thus, this high-throughput epigenomic strategy has served to identify novel hypermethylated candidates in bladder cancer. In vitro analyses supported the role of methylation in silencing SOX9 gene. The association of SOX9 hypermethylation with tumour progression and clinical outcome suggests its relevant clinical implications at stratifying patients affected with bladder cancer.

Gene expression profiles in HPV-infected head and neck cancer.

Epidemiological and laboratory evidence indicate that, in addition to tobacco and alcohol, human papillomaviruses (HPV) play an important aetiological role in a subset of head and neck squamous cell carcinoma (HNSCC). To evaluate the molecular pathogenesis of HPV-infected HNSCC, we compared gene expression patterns between HPV-positive and -negative HNSCC tumours using cDNA microarrays. Tumour tissue was collected from 42 histologically confirmed HNSCC patients from an inner-city area of New York. Total DNA and RNA were extracted and purified from frozen tumour samples and gene expression levels were compared to a universal human reference RNA standard using a 27 323 cDNA microarray chip. HPV detection and genotyping were performed using an MY09/11-PCR system and RT-PCR. HPV was detected in 29% of HNSCC tumours. Most harboured only HPV16 and expressed the HPV16-E6 oncogene. HPV prevalence was highest in pharyngeal tumours (45%). Gene expression patterns that differentiated HPV-positive from negative tumours were compared by supervised classification analysis, and a multiple-gene signature was found to predict HPV16 prevalence in primary HNSCC with a false discovery rate < 0.2. Focusing on never-smokers, we further identified a distinct subset of 123 genes that were specifically dysregulated in HPV16-positive HNSCC. Overexpressed genes in HPV-positive HNSCC tumours included the retinoblastoma-binding protein (p18), replication factor-C gene, and an E2F-dimerization partner transcription factor (TFDP2) that have also been found to be overexpressed in cervical cancer. An additional subset of genes involved in viral defence and immune response, including interleukins and interferon-induced proteins, was found to be down-regulated in HPV-positive tumours, supporting a characteristic and unique transcriptional profile in HPV-induced HNSCC.

Classification of DNA methylation patterns in tumor cell genomes using a CpG island microarray.

Our group has initiated experiments to epigenetically profile CpG island hypermethylation in genomic DNA from tissue specimens of head and neck squamous cell carcinoma (HNSCC) using a microarray of 12,288 CpG island clones. Our technique, known as a methylation-specific restriction enzyme (MSRE) analysis, is a variation of the differential methylation hybridization (DMH) technique, in that it is not an array comparison of two DNA samples using methylation-specific restriction enzymes. Instead, it is a comparison of a single DNA sample's response to a methylation-sensitive restriction enzyme (HpaII) and its corresponding methylation-insensitive isoschizomer (MspI). Estimation of the reproducibility of this microarray assay by intraclass correlation (ICC) demonstrated that in four replicate experiments for three tumor specimens, the ICC observed for a given tumor specimen ranged from 0.68 to 0.85 without filtering of data. Repeated assays achieved 87% concordance or greater for all tumors after filtering of array data by fluorescence intensity. We utilized hierarchical clustering on a population of 37 HNSCC samples to cluster tumor samples with similar DNA methylation profiles. Supervised learning techniques are now being utilized to allow us to identify associations between specific epigenetic signatures and clinical parameters. Such techniques will allow us to identify select groups of CpG island loci that could be used as epigenetic markers for both diagnosis and prognosis in HNSCC.