bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gastrostomy tube placement for long‐term oral drug administration in non‐human primates
Authors: Kim Jong‐Min , Shin Jun‐Seop , Min Byoung‐Hoon , Kim Jung‐Sik , Yoon Il‐Hee , Jeong Won‐Young , Lee Ga‐Eul , Kim Min‐Sun , Kim Ju‐Eun , Park Chung‐Gyu 
Published: March 2017

AbstractBackgroundNon‐human primates (NHPs) are often used as recipients in preclinical transplantation research that in most cases involves administration of various drugs including immunosuppressants. Long‐term oral drug administration, particularly tacrolimus, is challenging in the transplant recipient NHPs. Oral drug administration method using the mixture of drug and fruit juice has been used in NHPs, but this is not always effective in all monkeys. To those monkeys who are poorly compliant, oral drug administration in restraint or administration using gastrostomy tube should be necessary. The aim of this study was to compare the efficacy of between oral drug administration in restraint and administration using gastrostomy tube and to report complications and solutions to overcome the problems related to gastrostomy tube for long‐term oral drug dosing in rhesus monkeys.MethodsFifteen of 4‐ to 5‐year‐old male and female healthy rhesus monkeys weighing 5.0‐6.8 kg were used as recipients for porcine pancreatic islet transplantation. Oral drug administration in restraint was used for four monkeys, and gastrostomy tube was placed to other 11 monkeys (8‐French Feeding tube, n=6; Tri‐Funnel Replacement Gastrostomy tube, n=5). Oral immunosuppressive drugs such as sirolimus and tacrolimus were administered through the tube. The efficacy and the extent of ease for administration and related complications were compared between two groups.Results and ConclusionsThe complication of gastrostomy included a transient inflammation in the skin and peritonitis caused by a leakage around implantation site (one case), which could be overcome by changing suture method and tube type to interlocking box suture and Tri‐Funnel Replacement Gastrostomy tube, respectively. Despite these complications, oral drug administration using gastrostomy tube allowed us to perform accurate dosage of drug administration and to reduce the stress that both the monkey and the researcher may experience. Taken together, this study showed that gastrostomy tube placement is a better alternative to oral drug administration in restraint for long‐term oral drug administration in rhesus monkeys who tend to refuse to eat the mixture of drug and fruit juice.
Transgenic expression of human leukocyte antigen‐E attenuates GalKO.hCD46 porcine lung xenograft injury
Authors: Laird Christopher T. , Burdorf Lars , French Beth M. , Kubicki Natalia , Cheng Xiangfei , Braileanu Gheorghe , Sun Wenji , O'Neill Natalie A. , Cimeno Arielle , Parsell Dawn , So Edward , Bähr Andrea , Klymiuk Nikolai , Phelps Carol J. , Ayares David , Azimzadeh Agnes M. , Pierson Richard N. 
Published: March 2017

AbstractBackgroundLung xenografts remain susceptible to loss of vascular barrier function within hours in spite of significant incremental advances based on genetic engineering to remove the Gal 1,3‐αGal antigen (GalTKO) and express human membrane cofactor protein (hCD46). Natural killer cells rapidly disappear from the blood during perfusion of GalTKO.hCD46 porcine lungs with human blood and presumably are sequestered within the lung vasculature. Here we asked whether porcine expression of the human NK cell inhibitory ligand HLA‐E and β2 microglobulin inhibits GalTKO.hCD46 pig cell injury or prolongs lung function in two preclinical perfusion models.MethodsLungs from pigs modified to express GalTKO.hCD46 (n=37) and GalTKO.hCD46.HLA‐E (n=5) were harvested and perfused with human blood until failure or elective termination at 4 hours. Airway pressures and pulmonary artery hemodynamics were recorded in real time. Blood samples were also collected throughout the experiment for analysis. Porcine aortic endothelial cells (PAECs) from each genotype were cultured in monolayers in microfluidic channels and used in fluorescent cytotoxicity assays using human NK cells.ResultsHLA‐E expression on GalTKO.hCD46 PAECs was associated with significantly decreased antibody‐dependent and antibody‐independent NK‐mediated cytotoxicity under in vitro conditions simulating physiologic shear stress. Relative to GalTKO.hCD46 pig lungs perfused with human blood on an ex vivo platform, additional expression of HLA‐E increased median lung survival (>4 hours, vs 162 minutes, P=.012), and was associated with attenuated rise in pulmonary vascular resistance, and decreased platelet activation and histamine elaboration. As expected, HLA‐E expression was not associated with a significant difference in NK cell adhesion to endothelial cells in vitro, or NK cell and neutrophil sequestration during organ perfusion.ConclusionsWe conclude human NK cell activation contributes significantly to GalTKO.hCD46 pig endothelial injury and lung inflammation and show that expression of HLA‐E is associated with physiologically meaningful protection of GalTKO.hCD46 cells and organs exposed to human blood.
Biosynthesis of 2′-Chloropentostatin and 2′-Amino-2′-Deoxyadenosine Highlights a Single Gene Cluster Responsible for Two Independent Pathways in
Actinomadura
sp. Strain ATCC 39365
Authors: Gao Yaojie , Xu Gudan , Wu Pan , Liu Jin , Cai You-sheng , Deng Zixin , Chen Wenqing 
Published: March 2017

ABSTRACT

2′-Chloropentostatin (2′-Cl PTN, 2′-chloro-2′-deoxycoformycin) and 2′-amino-2′-deoxyadenosine (2′-amino dA) are two adenosine-derived nucleoside antibiotics coproduced by
Actinomadura
sp. strain ATCC 39365. 2′-Cl PTN is a potent adenosine deaminase (ADA) inhibitor featuring an intriguing 1,3-diazepine ring, as well as a chlorination at C-2′ of ribose, and 2′-amino dA is an adenosine analog showing bioactivity against RNA-type virus infection. However, the biosynthetic logic of them has remained poorly understood. Here, we report the identification of a single gene cluster (
ada
) essential for the biosynthesis of 2′-Cl PTN and 2′-amino dA. Further systematic genetic investigations suggest that 2′-Cl PTN and 2′-amino dA are biosynthesized by independent pathways. Moreover, we provide evidence that a predicted cation/H
+
antiporter, AdaE, is involved in the chlorination step during 2′-Cl PTN biosynthesis. Notably, we demonstrate that 2′-amino dA biosynthesis is initiated by a Nudix hydrolase, AdaJ, catalyzing the hydrolysis of ATP. Finally, we reveal that the host ADA (designated ADA1), capable of converting adenosine/2′-amino dA to inosine/2′-amino dI, is not very sensitive to the powerful ADA inhibitor pentostatin. These findings provide a basis for the further rational pathway engineering of 2′-Cl PTN and 2′-amino dA production.

IMPORTANCE
2′-Cl PTN/PTN and 2′-amino dA have captivated the great interests of scientists, owing to their unusual chemical structures and remarkable bioactivities. However, the precise logic for their biosynthesis has been elusive for decades. Actually, the identification and elucidation of their biosynthetic pathways not only enrich the biochemical repertoire of novel enzymatic reactions but may also lay solid foundations for the pathway engineering and combinatorial biosynthesis of this family of purine nucleoside antibiotics to generate novel hybrid analogs with improved features.
Pseudomonas aeruginosa Aggregate Formation in an Alginate Bead Model System Exhibits
In Vivo
-Like Characteristics
Authors: Sønderholm Majken , Kragh Kasper Nørskov , Koren Klaus , Jakobsen Tim Holm , Darch Sophie E. , Alhede Maria , Jensen Peter Østrup , Whiteley Marvin , Kühl Michael , Bjarnsholt Thomas 
Published: March 2017

ABSTRACT

Alginate beads represent a simple and highly reproducible
in vitro
model system for diffusion-limited bacterial growth. In this study, alginate beads were inoculated with
Pseudomonas aeruginosa
and followed for up to 72 h. Confocal microscopy revealed that
P. aeruginosa
formed dense clusters similar in size to
in vivo
aggregates observed
ex vivo
in cystic fibrosis lungs and chronic wounds. Bacterial aggregates primarily grew in the bead periphery and decreased in size and abundance toward the center of the bead. Microsensor measurements showed that the O
2
concentration decreased rapidly and reached anoxia ∼100 μm below the alginate bead surface. This gradient was relieved in beads supplemented with NO
3
−
as an alternative electron acceptor allowing for deeper growth into the beads. A comparison of gene expression profiles between planktonic and alginate-encapsulated
P. aeruginosa
confirmed that the bacteria experienced hypoxic and anoxic growth conditions. Furthermore, alginate-encapsulated
P. aeruginosa
exhibited a lower respiration rate than the planktonic counterpart and showed a high tolerance toward antibiotics. The inoculation and growth of
P. aeruginosa
in alginate beads represent a simple and flexible
in vivo
-like biofilm model system, wherein bacterial growth exhibits central features of
in vivo
biofilms. This was observed by the formation of small cell aggregates in a secondary matrix with O
2
-limited growth, which was alleviated by the addition of NO
3
−
as an alternative electron acceptor, and by reduced respiration rates, as well as an enhanced tolerance to antibiotic treatment.

IMPORTANCE
Pseudomonas aeruginosa
has been studied intensively for decades due to its involvement in chronic infections, such as cystic fibrosis and chronic wounds, where it forms biofilms. Much research has been dedicated to biofilm formation on surfaces; however, in chronic infections, most biofilms form small aggregates of cells not attached to a surface, but embedded in host material. In this study, bacteria were encapsulated in small alginate beads and formed aggregates similar to what is observed in chronic bacterial infections. Our findings show that aggregates are exposed to steep oxygen gradients, with zones of oxygen depletion, and that nitrate may serve as an alternative to oxygen, enabling growth in oxygen-depleted zones. This is important, as slow growth under low-oxygen conditions may render the bacteria tolerant toward antibiotics. This model provides an alternative to surface biofilm models and adds to the comprehension that biofilms do not depend on a surface for formation.
Construction of Bacillus thuringiensis Simulant Strains Suitable for Environmental Release
Authors: Park Sangjin , Kim Changhwan , Lee Daesang , Song Dong Hyun , Cheon Ki Cheol , Lee Hong Suk , Kim Seong Joo , Kim Jee Cheon , Lee Sang Yup 
Published: March 2017

ABSTRACT

For a surrogate bacterium to be used in outdoor studies, it is important to consider environmental and human safety and ease of detection. Recently,
Bacillus thuringiensis
, a popular bioinsecticide bacterium, has been gaining attention as a surrogate bacterium for use in biodefense. In this study, we constructed simulant strains of
B. thuringiensis
with enhanced characteristics for environmental studies. Through transposon mutagenesis, pigment genes were inserted into the chromosome, producing yellow-colored colonies for easy detection. To prevent persistence of spores in the environment, a genetic circuit was designed to produce a spore without sporulation capability. Two
loxP
sites were inserted, one on each side of the
spo0A
gene, which encodes a sporulation master regulator, and a sporulation-dependent Cre expression cassette was inserted into the chromosome. This genetic circuit successfully deleted
spo0A
during sporulation, producing spores that lacked the
spo0A
gene. In addition, two major α/β-type small acid-soluble spore protein (SASP) genes, predicted by synteny analysis, were deleted. The spores of the mutant strain showed increased UV-C sensitivity and quickly lost viability when tested in a solar simulator. When the spores of the mutant strain were administered to the lungs of BALB/c mice, cells were quickly removed from the body, suggesting enhanced
in vivo
safety. All strains constructed in this study contain no antibiotic resistance markers and all heterologous genes were inserted into the chromosome, which are useful features for simulants to be released into the environment.

IMPORTANCE
B. thuringiensis
has recently been receiving increasing attention as a good spore simulant in biodefense research. However, few studies were done to properly address many important features of
B. thuringiensis
as a simulant in environmental studies. Since spores can persist in the environment for years after release, environmental contamination is a big problem, especially when genetically engineered strains are used. To solve these problems, we report here the development of
B. thuringiensis
simulant strains that are capable of forming yellow colonies for easy detection, incapable of forming spores more than once due to a genetic circuit, and lacking in two major SASP genes. The genetic circuit to produce a spore without sporulation capability, together with the deletion of SASP genes, ensures the environmental and human safety of the simulant strains developed in this study. All of these features will allow wider use of
B. thuringiensis
as a simulant for
Bacillus anthracis
in environmental release studies.
Use of Natural Transformation To Establish an Easy Knockout Method in Riemerella anatipestifer
Authors: Liu MaFeng , Zhang Li , Huang Li , Biville Francis , Zhu DeKang , Wang MingShu , Jia RenYong , Chen Shun , Sun KunFeng , Yang Qiao , Wu Ying , Chen XiaoYue , Cheng AnChun 
Published: March 2017

ABSTRACT

Riemerella anatipestifer
is a member of the family
Flavobacteriaceae
and a major causative agent of duck serositis. Little is known about its genetics and pathogenesis. Several bacteria are competent for natural transformation; however, whether
R. anatipestifer
is also competent for natural transformation has not been investigated. Here, we showed that
R. anatipestifer
strain ATCC 11845 can uptake the chromosomal DNA of
R. anatipestifer
strain RA-CH-1 in all growth phases. Subsequently, a natural transformation-based knockout method was established for
R. anatipestifer
ATCC 11845. Targeted mutagenesis gave transformation frequencies of ∼10
−5
transformants. Competition assay experiments showed that
R. anatipestifer
ATCC 11845 preferentially took up its own DNA rather than heterogeneous DNA, such as
Escherichia coli
DNA. Transformation was less efficient with the shuttle plasmid pLMF03 (transformation frequencies of ∼10
−9
transformants). However, the efficiency of transformation was increased approximately 100-fold using pLMF03 derivatives containing
R. anatipestifer
DNA fragments (transformation frequencies of ∼10
−7
transformants). Finally, we found that the
R. anatipestifer
RA-CH-1 strain was also naturally transformable, suggesting that natural competence is widely applicable for this species. The findings described here provide important tools for the genetic manipulation of
R. anatipestifer
.

IMPORTANCE
Riemerella anatipestifer
is an important duck pathogen that belongs to the family
Flavobacteriaceae
. At least 21 different serotypes have been identified. Genetic diversity has been demonstrated among these serotypes. The genetic and pathogenic mechanisms of
R. anatipestifer
remain largely unknown because no genetic tools are available for this bacterium. At present, natural transformation has been found in some bacteria but not in
R. anatipestifer
. For the first time, we showed that natural transformation occurred in
R. anatipestifer
ATCC 11845 and
R. anatipestifer
RA-CH-1. Then, we established an easy gene knockout method in
R. anatipestifer
based on natural transformation. This information is important for further studies of the genetic diversity and pathogenesis in
R. anatipestifer
.
Prevalence, Genotype Richness, and Coinfection Patterns of Hemotropic Mycoplasmas in Raccoons (Procyon lotor) on Environmentally Protected and Urbanized Barrier Islands
Authors: Volokhov Dmitriy V. , Hwang Jusun , Chizhikov Vladimir E. , Danaceau Heather , Gottdenker Nicole L. 
Published: March 2017

ABSTRACTRaccoons (Procyon lotor) are successful urban adapters and hosts to a number of zoonotic and nonzoonotic pathogens, yet little is known about their hemoplasma infections and how prevalence varies across habitat types. This study identifies hemotropicMycoplasmaspecies infection in raccoons from urban and undisturbed habitats and compares hemoplasma infection in sympatric urban cats (Felis catus) from the same geographic region. We collected blood from raccoons (n= 95) on an urban coastal island (n= 37) and an undisturbed coastal island (n= 58) and from sympatric urban cats (n= 39) in Georgia, USA. Based on 16S rRNA gene amplification, 62.1% (59/95) of raccoons and 17.9% (7/39) of feral cats were positive for hemoplasma. There was a greater percentage of hemoplasma-infected raccoons on the undisturbed island (79.3% [46/58]) than on the urban island (35.1% [13/37]; χ2= 16.9,df= 1,P= 0.00004). Sequencing of the full-length 16S rRNA gene amplicons revealed six hemoplasma genotypes in raccoons, including five novel genotypes that were distinct from three known hemoplasma species identified in the sympatric cats. In addition, the hemoplasma genotypes detected in raccoons were not identified in sympatric cats or vice versa. Although all six hemoplasma genotypes were found in raccoons from urban and undisturbed islands, coinfection patterns differed between sites and among individuals, with the proportion of coinfected raccoons being greater in the undisturbed site. This study shows that raccoons are hosts for several novel hemoplasmas and that habitat type influences infection patterns.IMPORTANCEThis study provides information about novel hemoplasmas identified in raccoons (Procyon lotor), which can be used for assessments of the prevalence of these hemoplasmas in raccoon populations and for future studies on the potential pathogenic impacts of these hemoplasmas on raccoon health. Raccoons from the undisturbed habitat had a higher prevalence of hemoplasma infection than urban raccoons. There does not appear to be cross-species transmission of hemotropic mycoplasmas between urban raccoons and feral cats. Raccoons appear to be hosts for several novel hemoplasmas, and habitat type influences infection patterns.
Metatranscriptomic Evidence for Direct Interspecies Electron Transfer between Geobacter and Methanothrix Species in Methanogenic Rice Paddy Soils
Authors: Holmes Dawn E. , Shrestha Pravin M. , Walker David J. F. , Dang Yan , Nevin Kelly P. , Woodard Trevor L. , Lovley Derek R. 
Published: March 2017

ABSTRACTThe possibility thatMethanothrix(formerlyMethanosaeta) andGeobacterspecies cooperate via direct interspecies electron transfer (DIET) in terrestrial methanogenic environments was investigated in rice paddy soils. Genes with high sequence similarity to the gene for the PilA pilin monomer of the electrically conductive pili (e-pili) ofGeobacter sulfurreducensaccounted for over half of the PilA gene sequences in metagenomic libraries and 42% of the mRNA transcripts in RNA sequencing (RNA-seq) libraries. This abundance of e-pilin genes and transcripts is significant because e-pili can serve as conduits for DIET. Most of the e-pilin genes and transcripts were affiliated withGeobacterspecies, but sequences most closely related to putative e-pilin genes from genera such asDesulfobacterium,Deferribacter,Geoalkalibacter, andDesulfobacula, were also detected. Approximately 17% of all metagenomic and metatranscriptomic bacterial sequences clustered withGeobacterspecies, and the finding thatGeobacterspp. were actively transcribing growth-related genes indicated that they were metabolically active in the soils. Genes coding for e-pilin were among the most highly transcribedGeobactergenes. In addition, homologs of genes encoding OmcS, ac-type cytochrome associated with the e-pili ofG. sulfurreducensand required for DIET, were also highly expressed in the soils.Methanothrixspecies in the soils highly expressed genes for enzymes involved in the reduction of carbon dioxide to methane. DIET is the only electron donor known to support CO2reduction inMethanothrix. Thus, these results are consistent with a model in whichGeobacterspecies were providing electrons toMethanothrixspecies for methane production through electrical connections of e-pili.IMPORTANCEMethanothrixspecies are some of the most important microbial contributors to global methane production, but surprisingly little is known about their physiology and ecology. The possibility that DIET is a source of electrons forMethanothrixin methanogenic rice paddy soils is important because it demonstrates that the contribution thatMethanothrixmakes to methane production in terrestrial environments may extend beyond the conversion of acetate to methane. Furthermore, defined coculture studies have suggested that whenMethanothrixspecies receive some of their energy from DIET, they grow faster than when acetate is their sole energy source. Thus,Methanothrixgrowth and metabolism in methanogenic soils may be faster and more robust than generally considered. The results also suggest that the reason thatGeobacterspecies are repeatedly found to be among the most metabolically active microorganisms in methanogenic soils is that they grow syntrophically in cooperation withMethanothrixspp., and possibly other methanogens, via DIET.
Comparison of Fecal Collection Methods for Microbiota Studies in Bangladesh
Authors: Vogtmann Emily , Chen Jun , Kibriya Muhammad G. , Chen Yu , Islam Tariqul , Eunes Mahbubul , Ahmed Alauddin , Naher Jabun , Rahman Anisur , Amir Amnon , Shi Jianxin , Abnet Christian C. , Nelson Heidi , Knight Rob , Chia Nicholas , Ahsan Habibul , Sinha Rashmi 
Published: March 2017

ABSTRACT
To our knowledge, fecal microbiota collection methods have not been evaluated in low- and middle-income countries. Therefore, we evaluated five different fecal sample collection methods for technical reproducibility, stability, and accuracy within the Health Effects of Arsenic Longitudinal Study (HEALS) in Bangladesh. Fifty participants from the HEALS provided fecal samples in the clinic which were aliquoted into no solution, 95% ethanol, RNAlater, postdevelopment fecal occult blood test (FOBT) cards, and fecal immunochemical test (FIT) tubes. Half of the aliquots were frozen immediately at −80°C (day 0) and the remaining samples were left at ambient temperature for 96 h and then frozen (day 4). Intraclass correlation coefficients (ICC) were calculated for the relative abundances of the top three phyla, for two alpha diversity measures, and for four beta diversity measures. The duplicate samples had relatively high ICCs for technical reproducibility at day 0 and day 4 (range, 0.79 to 0.99). The FOBT card and samples preserved in RNAlater and 95% ethanol had the highest ICCs for stability over 4 days. The FIT tube had lower stability measures overall. In comparison to the “gold standard” method using immediately frozen fecal samples with no solution, the ICCs for many of the microbial metrics were low, but the rank order appeared to be preserved as seen by the Spearman correlation. The FOBT cards, 95% ethanol, and RNAlater were effective fecal preservatives. These fecal collection methods are optimal for future cohort studies, particularly in low- and middle-income countries.

IMPORTANCE
The collection of fecal samples in prospective cohort studies is essential to provide the opportunity to study the effect of the human microbiota on numerous health conditions. However, these collection methods have not been adequately tested in low- and middle-income countries. We present estimates of technical reproducibility, stability at ambient temperature for 4 days, and accuracy comparing a “gold standard” for fecal samples in no solution, 95% ethanol, RNAlater, postdevelopment fecal occult blood test cards, and fecal immunochemical test tubes in a study conducted in Bangladesh. Fecal occult blood test cards and fecal samples stored in 95% ethanol or RNAlater adequately preserve fecal samples in this setting. Therefore, new studies in low- and middle-income countries should include collection of fecal samples using fecal occult blood test cards, 95% ethanol, or RNAlater for prospective cohort studies.