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Advances in Microbiology 2017年第2期7卷 158-174页

作者： Shreyasee Chakraborty nwadiuto esiobu Microbial Biotech Laboratory Department of Biological Sciences Florida Atlantic University Boca Raton FL USA

The vital role of oral microbiome in the well-being of humans is only beginning to be unraveled. Employing a rigorous analysis of PCR-restriction fragment length polymorphisms (PCR-RFLP) and DNA fingerprints from denatured gradient gel electrophoresis (PCR-DGGE) of the 16S rDNA gene in metagenomic samples, this study evaluated the stability of the oral microbiome and contrasted the PCR-DGGE profiles of subjects belonging to three groups—healthy, smokers and oral diseases;in search of distinctive patterns predictive of each group. The DNA band size, intensity and profile generated by three restriction enzymes from a 1500 bp amplicon showed a fairly stable microbial community structure (P H' = 0.99 ± 0.12;S = 2.87 ± 0.75) and oral disease mouths (H' = 1.06 ± 0.085;S = 2.86 ± 0.31), than in healthy subjects (H' = 0.926 ± 0.07;S = 1.79 ± 0.56) suggesting that smoking is associated with a microbial community shift towards the structure found in poor oral health. It is clear that the human oral bacteria symbionts are not all random colonizers. Rather some of them constitute a fragile stable dynamic community whose disturbance could lead to disease or be indicative of disease. Understanding the dynamics of the bacterial community structure in health and disease states is a prerequisite to developing effective preventive healthcare and rapid diagnosis of diseases.

关键词： Oral Microbiome Metagenomic

Metagenomic Shotgun Sequencing Provides Prevalence Data for Pathogens, and Source-Tracking Indices Useful in Public Health Risk Assessment of Environmental Waters

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Journal of Geoscience and Environment Protection 2020年第6期8卷 115-129页

作者： Brian Mercer Karim Dawkins Lisa Meday nwadiuto esiobu Microbial Biotech Lab Sanson Building Florida Atlantic University Boca Raton USA Environmental Resource Management Department Water Quality Seminole Tribe of Florida Holywood USA

State-approved membrane filtration (MF) techniques for water quality assessments were contrasted with metagenomic shotgun sequencing (MSS) protocols to evaluate their efficacy in providing precise health-risk indices for surface waters. Samples from a freshwater receiving pond (ABI-1002) and two upstream storm water ditches (ABI-1003) and (ABI-1004) yielded alarmingly high Fecal coliform MF densities of 220, >2000 and >2000 CFU/100ml respectively. The indicator, Enterococcus bacteria exceeded allowable limits in all but the equipment control (ABI-1001). Using MSS, the relative numerical abundance of pathogenic bacteria, virulence and antibiotic resistance genes revealed the status and potential pollution sources of each ditch. High levels of Shigella sp. (0 (ABI-1001), 4945 (ABI-1002), 55,008 (ABI-1003), and 2221 (ABI-1004) genomic reads/100ml) correlated with virulence genes and antibiotic resistance genes found in fecal samples for ABI1003 and not ABI1004. Traditional culture methods (TCM) showed possible fecal contamination in two of the four samples, and no contamination in the others. MSS clearly distinguished between fecal and environmental bacteria contamination sources, and pinpointed actual risks from pathogens. Our data underscore the potential utility of MSS in precision risk assessment for public and biodiversity health and tracking of environmental microbiomes shifts by field managers and policy makers.

关键词： Metagenomic Shotgun Sequencing Water Quality Risk Assessment Indicator Bacteria Environmental Microbiomes

Analysis of the Human Oral Microbiome of Smokers and Non-Smokers Using PCR-RFLP and Ribotyping

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Advances in Microbiology 2014年第10期4卷 681-691页

作者： Shreyasee Chakraborty Verneshia Persaud Sonia Vanegas Gloryanne Gautier nwadiuto esiobu Department of Biological Sciences College of Sciences Florida Atlantic University at Davie Davie FL US Department of Biological Sciences College of Sciences Florida Atlantic University at Davie Davie FL US

Recent advances in the field of microbial and medical ecology emphasize the critical role played by oral bacteria in the delicate dynamic equilibrium of human health and disease, creating the need to define the bacterial communities associated with healthy and non-healthy conditions and to capture shifts in community structure germane to diagnosis. Employing PCR-RFLP of the 16S rDNA gene from metagenomes and plate-wash (cultured) bacteria of oral wash from 10 volunteers, this study evaluated the stability of oral bacteria in healthy subjects and documented community shifts in smokers. Sequence analysis of selected 16S gene amplicons cloned with the Gene Hunter PCR-Trap vector and pCR 4-TOPO cloning kits was conducted to determine the bacteria identity and diversity indices of the two groups. Ribopatterns generated by the restriction enzymes HaeIII and Sau3AI were significantly (p AluI using the GelCompare II software cluster analysis. A stable core of bacteria DNA fingerprint was detected in all healthy subjects, and remained unchanged over the study period of 3 months. Signature bands (1500 bp with HaeIII) in smokers and in non-smokers (800 bp and 700 bp with Sau3A1) were evidently suggesting the presence of potential biomarkers of healthy and non-healthy states. There was no significant difference in the DNA fingerprints of cultured and metagenomic extracts. The genera Xanthomonas, Streptococcus and phylum Candidatus occurred in large numbers in both groups, however, a major shift in composition with the dominance of gram-negative bacteria in smokers compared to healthy subjects was quite remarkable. Taxonomic diversity in smokers was quite high, including members of the genera Rothia, Synechococcus, Neisseria, Thiomargarita and Pyrobaculum. These data highlight the presence of a stable core microbiome amidst a wide diversity, identify a distinct smokers’ cluster and open the way for the search for potential biomarkers for specific diseases.

关键词： Oral Microbiome PCR-RFLP 16S rDNA Sequence Restriction Enzymes

Different Detection and Treatment Methods for <i>Entamoeba histolytica</i>and <i>Entamoeba dispar</i>in Water/Wastewater: A Review

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Journal of Environmental Protection 2022年第1期13卷 126-149页

作者： Rakib Ahmed Chowdhury nwadiuto esiobu Daniel E. Meeroff Fred Bloetscher Department of Civil Environmental & Geomatics Engineering Florida Atlantic University Boca Raton FL USA Department of Biological Sciences Florida Atlantic University Boca Raton FL USA

Entamoeba histolytica is an anaerobic parasitic protozoan and well known as a human pathogen, while its close relative, Entamoeba dispar, also possesses similar characteristics as an infectious agent. These microorganisms are generally transmitted in fecal-contaminated water. However, E. dispar present in industrial wastewater is also capable of creating biofilms that can cause adverse impacts in piping networks. Therefore, it is important to detect both of these protozoan species in water and to find a cost-effective technique for inactivation or management control. This review article summarizes the available detection methods in water and wastewater matrices along with feasible disinfection techniques.

关键词： Entamoeba histolytica Entamoeba dispar Detection Methods Water and Wastewater Treatment

Synthetic Biology Construct of Ebola Virus in Bacteria Surrogate Is Stable and Safe for Rapid Detection Studies in a BSL-2 Laboratory Setting

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Advances in Microbiology 2022年第1期12卷 25-41页

作者： nwadiuto esiobu Douglas Holmes Chad Coarsey Waseem Asghar Bodhi Stone Microbial Biotech Laboratory Biological Sciences Department Florida Atlantic University Boca Raton Florida USA Department of Electrical Engineering and Computer Science Florida Atlantic University Boca Raton Florida USA

Rapid detection of virulent pathogens during an outbreak is critical for public health advisories and control of the disease in a population. While many molecular techniques for point of care and clinical diagnosis abound, the US experience with the COVID-19 testing in the early stages of the pandemic underscores the critical importance of determining the appropriate target gene(s) with in-built controls that reliably detect pathogens with high sensitivity and specificity. Assays and research for diagnostics and therapy could be slowed during an epidemic because access to the required BSL-3 and BSL-4 laboratories are limited. So, during the 2014 West Africa Ebola outbreak, we tested the hypothesis that using synthetic cDNA of Ebolavirus in a bacteria surrogate (fit for all lab settings), would remain unmutated and safe after several generations, serving as an effective positive control in research settings, self test and point-of-care detection platforms. Primers were designed for the detection and quantification of the nucleoprotein (NP) gene of the 2014 Makona Ebola strain (KR781608.1, 733 - 1332 bp). To test the stability of artificially inserted translation arrest in the Orf of the model gene, it was edited to include three STOP codons in the RNA transcript using SNAP GENE. The segment was then spliced into a high copy number plasmid, cloned into One ShotTM TOP10 Escherichia coli (Invitrogen), and tested for stability and safety by periodic subculture, extraction and sequencing. Unlike COVID-19, rapid detection of blood-borne etiologies like Ebola requires optimized protocols for blood matrix. Using real-time PCR and newly designed primer pairs, the EBOV surrogate was detected and enumerated in human blood and regular broth and buffers. Based on aligned sequence analysis, the EBOV synthetic NP gene was stable (>99.9999% similarity coefficient) for at least 3 months. Detection sensitivity in broth and blood was at least 100 cells/ml or about 5.8

关键词： Ebola Virus Synthetic Biology Biosafety Point-of-Care Rapid Detection Plasmid Vector

Prevalence and Disinfection of Bacteria Associated with Various Types of Wristbands

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Advances in Infectious Diseases 2023年第2期13卷 193-209页

作者： Joseph Mendonca Belen Wertheimer Daynalee Dixon Bodhi Stone Karim Dawkins Miranda Christian nwadiuto esiobu Microbial Biotechnology Laboratory Department of Biological Sciences Florida Atlantic University Boca Raton FL USA West Palm Beach TV (WPTV) West Palm Beach FL USA

The potential role of personal items in the transmission of pathogens is poorly understood. In this study, we cultured bacteria of public health importance found on wristbands, determined whether there is a correlation between wristband material and prevalence, and tested three household disinfectants for efficacy in reducing bacteria on wristbands made of plastic, rubber, metal, and cloth, using standard microbiological assays. Total cultivable bacteria, Staphylococci, Enterobacteria (Escherichia coli), and Pseudomonas on 20 smartwatch wristbands were cultured from randomly recruited subjects. Nearly all wristbands (95%) were contaminated, with the highest average numbers of 3.46E+4 cfu/cm2 and 1.52E+4 cfu/cm2 on rubber and plastic bands respectively. Metallic gold and silver wristbands had zero to 18 cfu/cm2. While the high prevalence of Staphylococcus spp (85% of wristbands)—skin microbiota;was not unexpected, the occurrence of Pseudomonas spp (30%), and enteric bacteria (60%), even at relatively low numbers is of public health significance. Bacterial load on individual subjects varied remarkably with males and females harboring average total bacteria of 4.045 and 3.42 log10cfu/cm2 of wristband, respectively. The most important predictor of wristband bacteria load was the texture of wristband material and activity (hygiene) of the subject at sampling time. Potential pathogens—Staphylococcus aureus (8143 cfu/cm2) and Pseudomonas spp. (1126 cfu/cm2) were most abundant on cloth and rubber wristbands, respectively, while the presence of the E. coli group was associated with animal handling activity by a veterinarian. Lysol Disinfectant Spray and 70% Ethanol were highly effective regardless of wristband material with >99.99% kill rate and a log cfu/cm2 reduction of 3 - 4.0 and 3 - 4.5 respectively within 30 seconds. Apple Cider Vinegar (ACV) was not as potent. Only 2 - 3.5 log cfu/

关键词： Apple Cider Vinegar Disinfectants Escherichia coli Lysol Pseudomonas aeruginosa Public Health Staphylococcus aureus Wristbands

Analysis of a Tropical Warm Spring Microbiota Using 16S rRNA Metabarcoding

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Advances in Microbiology 2020年第4期10卷 145-165页

作者： Deborah Fasesan Karim Dawkins Roberto Ramirez Hadiza Rasheed-Jada Abiodun Onilude Oyekanmi Nash nwadiuto esiobu Center for Genomics Research and Innovation National Biotechnology Development Agency Abuja Nigeria Department of Microbiology University of Ibadan Ibadan Nigeria Microbial Biotechnology Laboratory Florida Atlantic University Boca Raton USA

The Ikogosi Warm Spring is a unique ecological niche in Western Nigeria with an average temperature and pH of 38°C and 5.8 respectively. It mixes with an adjacent cold spring (28°C & pH 7.6), about 100 meters from source, yielding a confluence body of water of 32°C and pH 7.7. To explore the bacterial community structure of this uncommon environment and to scan for potentially useful bacteria, metagenomes extracted directly from five samples (source and mid-point of warm spring;source and midpoint of cold spring, and the confluence) were analyzed. Using the MiSeq Illumina next generation sequencing protocols, the V3-V4 region of the 16S rRNA gene pool was sequenced and analyzed by QIIME (Quantitative Insights into Microbial Ecology) and R software. At least 11% (47,446) of all the sequences were unknown to any of the databases employed. Bacterial diversity and abundance at the source of both springs were extremely low, accounting for less than 0.07% of the total sequence reads at the confluence, 100 m downstream. In contrast to the highly diversified mesophilic confluence community where 21 different phyla were identified, only 4 and 5 phyla were recovered from the source-point of the warm spring and cold spring respectively. The most prevalent phyla in all samples were members of the versatile Proteobacteria (35% - 50% relative abundance), and the hardy Firmicutes (33% - 40%). Operational taxonomic units (OTUs) obtained from all the samples averaged at 1414. Temperature and pH were equally significant predictors of genomic diversity and richness, with the warm and cold spring sources having less than 5 bacteria phyla. Exiguobacterium sp. (a potential plastic degrader) and other deep rooted bacteria were found in the warm spring while the cold spring outflow contained among others such as Rubrobacter sp. and Chloroflexi sp. (which is close to the phylogenetic root of the domain Bacteria). Many taxonomically unresolved sequences could indicate the presence of potenti

关键词： Ikogosi Warm Springs Amplicon Metagenomics Next-Gen Sequencing QIIME Bacterial Diversity Source

Microbiomes of Top and Sub-Layers of Semi-Arid Soils in North-Eastern Nigeria Are Rich in Firmicutes and Proteobacteria with Surprisingly High Diversity of Rare Species

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Advances in Microbiology 2019年第1期9卷 102-118页

作者： Mwajim Bukar Oluwole Sodipo Karim Dawkins Roberto Ramirez Jummai T. Kaldapa Martha Tarfa nwadiuto esiobu North-East Zonal Biotechnology Centre of Excellence University of Maiduguri Maiduguri Nigeria Biochemistry Department Faculty of Science University of Maiduguri Maiduguri Nigeria Biological Sciences Department Microbial Biotech Laboratory Florida Atlantic University Boca Raton FL USA

Borno state is the second largest state in Nigeria with over 70,000 square kilometers of diverse ecosystems including parts of the fertile Lake Chad basin. However, more than 2/3 of this landmass is threatened with drought, advancing desertification and degraded soils. Most restoration efforts involve revegetation, which in the past has met with limited success. Microbial communities of soils play a pivotal role in soil fertility and plant cover. We conducted the first metagenomic amplicon sequencing study, comparing two soil depths to determine whether soil bacteria abundance and diversity in the harsh bare soils were sufficient to sustain greening efforts. The goal was to glean insights to guide microbial inoculant formulation needed in the region. Samples from top (0 - 15 cm) and sub (16 - 65 cm) soils were collected from five strategic locations in the state. Using next generation Illumina sequencing protocols, total DNA extracted directly from the soils was sequenced and analyzed by QIIME. Metadata collected from site showed scorching temperatures of over 46?C, near zero moisture level and a pH of about 6 for top soil. At 65 cm depth, the temperature averaged 32?C with a pH of 5 and significantly higher soil moisture of 0.1%. The bacterial community structure was unexpectedly very diverse at both soil depths samples, recording a ChaO1 index ranging from 909 to 4296 and a Shannon diversity range of 3.54 to 6.33. The most abundant phyla in both soil depths were the Firmicutes and Proteobacteria;however the relative abundance of composite lower taxa was strikingly different. Operational taxonomic units and diversity indices were highest for top soils and were dominated by members of resilient groups of Actinobacteria, Firmucutes, Acidobacteria and numerous other less well-known taxa whose individual relative abundance did not exceed 3% of total population. The high diversity and richness of Proteobacteria (at 65 cm depth), some of which are key to soil fertility,

关键词： Microbiomes Semi-Arid Soils Microbial Community Structure Soil Bacteria Soil Fertility Top-Soil Re-Vegetation Proteobacteria Firmicutes Sub-Surface OTU

Characterization of Biofilms and Mineralogical Scale in Underground Injection Well Disposal of Landfill Leachate and Industrial Wastewater Streams

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Journal of Geoscience and Environment Protection 2019年第11期7卷 69-91页

作者： Daniel E. Meeroff Bishow Shaha Frederick Bloetscher nwadiuto esiobu Brian Mercer Donald McCorquordale Ramana Kari Michael Bennett Department of Civil Environmental & Geomatics Engineering Florida Atlantic University Boca Raton FL USA Department of Biological Sciences Florida Atlantic University Sanson Life Sciences Boca Raton FL USA Applied Biotech Inc. Plantation FL USA Solid Waste Authority of Palm Beach County West Palm Beach FL USA AECOM/URS West Palm Beach FL USA

Deep-well injection is a cost-effective alternative for industrial wastewater disposal, given the appropriate geology. Fouling of injection well tubing by biofilm or scale is common and reduces the effective diameter of the pipe, which results in increased wellhead pressure and lower injectivity. A detailed microbiological composition of biofilms and abiotic fingerprints use of mineral scale from an injection well has not been reported before. Therefore, biofilm and mineral scale samples from three depth intervals within a deep injection well (surface zone, D1= 0 - 61 m;intermediate zone, D2 = 62 - 457 m;and above the injection zone, D3 = 458 - 884 m) with recurrent biofilm development were collected to characterize the mineral composition and microbial community DNA. X-ray diffraction (XRD) analysis of the solids confirms the composition of the solids collected was mostly calcium carbonate (CaCO3) with minor contributions from common mineral salts. Microbiological fingerprinting suggests that methanogenic archaea and sulfate-reducing bacteria both of which are anaerobic biofilm producers were the most prevalent members of the prokaryotic community at all sampled depths. Methanosarcinae spp. increased with increasing depths, unlike other archaea. A non-pathogenic biofilm-producing Entamoeba dispar was the most prevalent member of the microbial domain (>30%) in all samples but was highest at the middle depth. The Chao alpha diversity indices for bacteria, viruses, and protozoans were highest at the shallow depth and gradually declined with increasing depth. The prevalent species above the injection zone depth are not barophilic organisms that thrive at high pressures, rather they are sulfate-reducing bacteria involved in anaerobic dissimilatory sulfate metabolism.

关键词： Injection Well Leachate Industrial Wastewater Disposal Biofilm Microbiology Well Rehabilitation Metagenomic Sequencing