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fecal DNA extraction

Rebekah C Kennedy, Russell R Fling, Michael S Robeson, Arnold M Saxton, Liesel G Schneider, John L Darcy, David A Bemis, Ling Zhao, Jiangang Chen
Widely used as an antimicrobial in antibacterial bar soaps, triclocarban (3,4,4'-trichlorocarbanilide; TCC) is effective against Gram-positive bacteria but shows little efficacy against Gram-negative strains, potentially altering the composition of indigenous microflora within and on the human body. To date, the consequence of continuous or previous nonprescription antimicrobial exposure from compounds in personal care products on commensal microflora is still elusive. Previous research has shown that TCC exposure during gestation and lactation induced dysbiosis of gut microbial communities among exposed dams and neonates...
March 13, 2018: Environmental Science and Pollution Research International
Jocelyn Sietsma Penington, Megan A S Penno, Katrina M Ngui, Nadim J Ajami, Alexandra J Roth-Schulze, Stephen A Wilcox, Esther Bandala-Sanchez, John M Wentworth, Simon C Barry, Cheryl Y Brown, Jennifer J Couper, Joseph F Petrosino, Anthony T Papenfuss, Leonard C Harrison
To optimise fecal sampling for reproducible analysis of the gut microbiome, we compared different methods of sample collection and sequencing of 16S rRNA genes at two centers. Samples collected from six individuals on three consecutive days were placed in commercial collection tubes (OMNIgeneGut OMR-200) or in sterile screw-top tubes in a home fridge or home freezer for 6-24 h, before transfer and storage at -80 °C. Replicate samples were shipped to centers in Australia and the USA for DNA extraction and sequencing by their respective PCR protocols, and analysed with the same bioinformatic pipeline...
March 12, 2018: Scientific Reports
Lisa F Stinson, Jeffrey A Keelan, Matthew S Payne
The establishment of human gut microbiota commences initially in utero . Meconium-the first fecal material passed after birth-can be used to study fetal gut contents; however, processing meconium samples for microbiome studies presents significant technical challenges. Meconium hosts a low biomass microbiome, is tar-like in texture and contains high concentrations of PCR inhibitors. This study aimed to evaluate four different DNA extraction methods to elucidate the most effective method for bacterial DNA recovery and sequencing analysis from first-pass meconium...
2018: Frontiers in Microbiology
Qian Zhang, Hongyue Yu, Xinhua Xiao, Ling Hu, Fengjiao Xin, Xiaobing Yu
Background & Aims: Accumulating research has addressed the linkage between the changes to gut microbiota structure and type 2 diabetes (T2D). Inulin is one type of soluble dietary fiber that can alleviate T2D. As a prebiotic, inulin cannot be digested by humans, but rather is digested by probiotics. However, whether inulin treatment can benefit the entire gut bacteria community remains unknown. In this study, we evaluated the differences in gut microbiota composition among diabetic, inulin-treated diabetic, normal control, and inulin-treated normal control rats...
2018: PeerJ
L Calamari, P Morera, P Bani, A Minuti, L Basiricò, A Vitali, U Bernabucci
High temperature influences rumen and gut health, passage rate, and diet digestibility, with effects on fermentative processes. The main aim of the study was to investigate the effect of hot season on hindgut fermentation, the occurrence of Clostridium tyrobutyricum spores in bovine feces, and on their relationship with metabolic conditions in dairy cows producing milk used for Grana Padano cheese. The study was carried out on 7 dairy farms located in the Po Valley (Italy), involving 1,950 Italian Friesian dairy cows...
February 28, 2018: Journal of Dairy Science
Bruna Nicoleti Santana, Brayan Kurahara, Alex Akira Nakamura, Vinícius da Silva Camargo, Elis Domingos Ferrari, Giane Serafim da Silva, Walter Bertequini Nagata, Marcelo Vasconcelos Meireles
The objective of this study was to determine the occurrence of Cryptosporidium spp. in domestic chickens raised in different chicken production systems in Brazil using three nested PCR protocols. The purification and concentration of oocysts present in 190 fecal samples from chickens raised in extensive, semi-intensive and intensive production systems were accomplished by centrifugal flotation in Sheather's solution and were followed by the extraction of genomic DNA. The detection and molecular characterization of Cryptosporidium species and genotypes were performed using three nested polymerase chain reaction (nested PCR) protocols targeting the 18S rRNA gene followed by sequencing of the amplified fragments...
March 1, 2018: Preventive Veterinary Medicine
Wei-Kai Wu, Chieh-Chang Chen, Suraphan Panyod, Rou-An Chen, Ming-Shiang Wu, Lee-Yan Sheen, Shan-Chwen Chang
Although great interest has been displayed by researchers in the contribution of gut microbiota to human health, there is still no standard protocol with consensus to guarantee the sample quality of metagenomic analysis. Here we reviewed existing methodology studies and present suggestions for optimizing research pipeline from fecal sample collection to DNA extraction. First, we discuss strategies of clinical metadata collection as common confounders for microbiome research. Second, we propose general principles for freshly collected fecal sample and its storage and share a DIY stool collection kit protocol based on the manual procedure of Human Microbiome Project (HMP)...
February 25, 2018: Journal of the Formosan Medical Association, Taiwan Yi Zhi
Liu Xiao-Jie, Mao Tie-Bo, Wu Peng, Zhou Rui
OBJECTIVE: To understand the prevalence of Cryptosporidium infection in diarrhea infants under 2 years old in Wuhan City, so as to provide the epidemiological evidence for the prevention and treatment of cryptosporidiosis. METHODS: The fecal samples from infants under 2 years old with diarrhea were collected in Hubei General Hospital and Central South Hospital in Wuhan City, Hubei Province from August 2014 to July 2015. The fecal samples were stored in 2.5% potassium dichromate at 4 ℃ after filtered...
March 14, 2017: Zhongguo Xue Xi Chong Bing Fang Zhi za Zhi, Chinese Journal of Schistosomiasis Control
Nicholas A Kennedy, Christopher A Lamb, Susan H Berry, Alan W Walker, John Mansfield, Miles Parkes, Rachel Simpkins, Mark Tremelling, Sarah Nutland, Julian Parkhill, Chris Probert, Georgina L Hold, Charlie W Lees
Background/Aims: Current models of Crohn's disease (CD) describe an inappropriate immune response to gut microbiota in genetically susceptible individuals. NOD2 variants are strongly associated with development of CD, and NOD2 is part of the innate immune response to bacteria. This study aimed to identify differences in fecal microbiota in CD patients and non-IBD controls stratified by NOD2 genotype. Methods: Patients with CD and non-IBD controls of known NOD2 genotype were identified from patients in previous UK IBD genetics studies and the Cambridge bioresource (genotyped/phenotyped volunteers)...
February 15, 2018: Inflammatory Bowel Diseases
Timothy J Divoll, Veronica A Brown, Jeff Kinne, Gary F McCracken, Joy M O'Keefe
Different second-generation sequencing technologies may have taxon-specific biases when DNA metabarcoding prey in predator feces. Our major objective was to examine differences in prey recovery from bat guano across two different sequencing workflows using the same fecal DNA extracts. We compared results between the Ion Torrent PGM and the Illumina MiSeq with similar library preparations and the same analysis pipeline. We focus on repeatability and provide an R Notebook in an effort toward transparency for future methodological improvements...
February 17, 2018: Molecular Ecology Resources
Djursun Karasartova, Ayse Semra Gureser, Emrah Ruh, Buse Turegun-Atasoy, Mustafa Kerem Calgin, Leyla Tasci, Aysegul Taylan-Ozkan
Polymerase chain reaction (PCR) is an effective technique for diagnosis of Blastocystis infection. Notably, DNA isolation procedure is extremely critical for the PCR step. In the present study, a recently described extraction procedure, named as the "sand method" was modified and adapted for isolation of Blastocystis DNA. To evaluate its efficacy, the current method and QIAamp DNA Stool Mini Kit (Qiagen) were applied to fresh human stool samples. Our results indicated that, the mean DNA concentrations obtained by the sand method and the commercial kit were 48 and 55 ng/μl, respectively...
February 10, 2018: Experimental Parasitology
Shin Hamada, Atsushi Masamune, Tatsuhide Nabeshima, Tooru Shimosegawa
Host-derived factors alter gut microenvironment, and changes in gut microbiota also affect biological functions of host. Alterations of gut microbiota have been reported in a wide variety of diseases, but the whole picture of alterations in pancreatic diseases remains to be clarified. In particular, the gut microbiota may be affected by malnutrition or impaired exocrine pancreas function that is associated with pancreatic diseases. We here conducted comprehensive analysis of gut microbiota in patients with type 1 autoimmune pancreatitis (AIP), a pancreatic manifestation of the systemic IgG4-related disease, and chronic pancreatitis (CP)...
2018: Tohoku Journal of Experimental Medicine
Călin Mircea Gherman, Zsuzsa Kalmár, Adriana Györke, Viorica Mircean
BACKGROUND: Giardia duodenalis is a parasitic protist that infects a large number of species, being localized in the small intestine. Two of the eight recognized assemblages have zoonotic potential, but studies regarding their distribution in less important pet or farm species are scarce. Of these species, the long-tailed chinchilla is a host for Giardia spp., although data on the spread of infection and assemblages involved are confined. The present work aimed to determine the prevalence of Giardia infection and assemblage identification in farmed chinchillas in Romania...
February 7, 2018: Parasites & Vectors
Abdollah Derakhshandeh, Vida Eraghi, Azar Motamedi Boroojeni, Malihe Akbarzadeh Niaki, Sahar Zare, Zahra Naziri
Escherichia coli (E. coli) is a normal flora of gastrointestinal tracts of humans and warm-blooded animals including dogs that has close vicinity with humans. Because the inter-species transmission of E. coli between pets and human beings, within a household, obtaining more information about the epidemiology, genetics, virulence factors, and antibiotic resistance of E. coli from dogs and their owners will help to control the inter-species transmission and treatment of E. coli infections. In this study we characterize and compare the antibiotic resistance and virulence profiles of fecal E...
January 30, 2018: Microbial Pathogenesis
Ibrahim Hamad, Rita Abou Abdallah, Isabelle Ravaux, Saadia Mokhtari, Hervé Tissot-Dupont, Caroline Michelle, Andreas Stein, Jean-Christophe Lagier, Didier Raoult, Fadi Bittar
Research on the relationship between changes in the gut microbiota and human disease, including AIDS, is a growing field. However, studies on the eukaryotic component of the intestinal microbiota have just begun and have not yet been conducted in HIV-infected patients. Moreover, eukaryotic community profiling is influenced by the use of different methodologies at each step of culture-independent techniques. Herein, initially, four DNA extraction protocols were compared to test the efficiency of each method in recovering eukaryotic DNA from fecal samples...
2018: PloS One
L Martinez-Anton, M Marenda, S M Firestone, R N Bushell, G Child, A I Hamilton, S N Long, M A R Le Chevoir
BACKGROUND: Acute polyradiculoneuritis (APN) is an immune-mediated peripheral nerve disorder in dogs that shares many similarities with Guillain-Barré syndrome (GBS) in humans, in which the bacterial pathogen Campylobacter spp. now is considered to be a major triggering agent. Little information is available concerning the relationship between APN and Campylobacter spp. in dogs. HYPOTHESIS/OBJECTIVES: To estimate the association between Campylobacter spp. infection and APN...
January 22, 2018: Journal of Veterinary Internal Medicine
Marina Papaiakovou, Nils Pilotte, Ben Baumer, Jessica Grant, Kristjana Asbjornsdottir, Fabian Schaer, Yan Hu, Raffi Aroian, Judd Walson, Steven A Williams
BACKGROUND: Proper collection and storage of fecal samples is necessary to guarantee the subsequent reliability of DNA-based soil-transmitted helminth diagnostic procedures. Previous research has examined various methods to preserve fecal samples for subsequent microscopic analysis or for subsequent determination of overall DNA yields obtained following DNA extraction. However, only limited research has focused on the preservation of soil-transmitted helminth DNA in stool samples stored at ambient temperature or maintained in a cold chain for extended periods of time...
January 2018: PLoS Neglected Tropical Diseases
Andrew Scott, Yuan-Ching Tien, Craig F Drury, W Daniel Reynolds, Edward Topp
The impact of amendment with swine manure composts (SMC), yard waste compost (YWC), or food waste compost (FWC) on the abundance of antibiotic resistance genes in soil was evaluated. Following a commercial-scale application of the composts in a field experiment, soils were sampled periodically for a decade, and archived air-dried. Soil DNA was extracted and gene targets quantified by qPCR. Compared to untreated control soil, all three amendment types increased the abundance of gene targets for up to four years post-application...
January 17, 2018: Canadian Journal of Microbiology
Mohammad Rayani, Gholamreza Hatam, Ngah Zasmy Unyah, Abdolmajid Ashrafmansori, Wan Omar Abdullah, Rukman Awang Hamat
Background: This study is the first phylogenetic genotype analysis of Giardia lamblia in Iran. The main objective was to determine genotyping and identify the sub-assemblages of Giardia lamblia isolates involved in the transmission of giardiasis in Fars Province, south of Iran, in 2012. Methods: Forty G. lamblia isolates were collected from the patient's fecal samples with gastrointestinal discomfort referred to the health centers and hospitals in Shiraz, Fars Province, south of Iran...
October 2017: Iranian Journal of Parasitology
Mi Young Lim, Eun-Ji Song, Sang Ho Kim, Jangwon Lee, Young-Do Nam
The human gut harbors a vast range of microbes that have significant impact on health and disease. Therefore, gut microbiome profiling holds promise for use in early diagnosis and precision medicine development. Accurate profiling of the highly complex gut microbiome requires DNA extraction methods that provide sufficient coverage of the original community as well as adequate quality and quantity. We tested nine different DNA extraction methods using three commercial kits (TianLong Stool DNA/RNA Extraction Kit (TS), QIAamp DNA Stool Mini Kit (QS), and QIAamp PowerFecal DNA Kit (QP)) with or without additional bead-beating step using manual or automated methods and compared them in terms of DNA extraction ability from human fecal sample...
December 21, 2017: Systematic and Applied Microbiology
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