Aptamers are short single-stranded DNA or RNA oligonucleotides
capable of binding with high affinity and specificity
to target molecules. Because of their durability and ease of synthesis,
aptamers are used in a wide range of biomedical fields,
including the diagnosis of diseases and targeted delivery of
therapeutic agents. The aptamers were selected using a process
called systematic evolution of ligands by exponential enrichment
(SELEX), which has been improved for various research
purposes since its development in 1990. In this protocol,
we describe a modified SELEX method that rapidly produces
high aptamer screening yields using two types of magnetic
beads. Using this method, we isolated an aptamer that
specifically binds to an antimicrobial peptide. We suggest that
by conjugating a small therapeutic-specific aptamer to a gold
nanoparticle-based delivery system, which enhances the stability
and intracellular delivery of peptides, aptamers selected
by our method can be used for the development of therapeutic
agents utilizing small therapeutic peptides.
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Recent approaches in the application of antimicrobial peptides in food preservation Satparkash Singh, Bhavna Jha, Pratiksha Tiwari, Vinay G. Joshi, Adarsh Mishra, Yashpal Singh Malik World Journal of Microbiology and Biotechnology.2024;[Epub] CrossRef
Design and application of microfluidics in aptamer SELEX and Aptasensors Shikun Zhang, Yingming Zhang, Zhiyuan Ning, Mengxia Duan, Xianfeng Lin, Nuo Duan, Zhouping Wang, Shijia Wu Biotechnology Advances.2024; 77: 108461. CrossRef
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Two novel Gram-positive, non-spore-forming, facultatively
anaerobic, non-motile, and short rods to coccoid strains were
isolated from the feces of the greater white-fronted geese
(Anser albifrons) at Poyang Lake. The 16S rRNA gene sequences
of strains 4H37-19T and 3HC-13 shared highest identity
to that of Corynebacterium uropygiale Iso10T (97.8%).
Phylogenetic and phylogenomic analyses indicated that strains
4H37-19T and 3HC-13 formed an independent clade within
genus Corynebacterium and clustered with Corynebacterium
uropygiale Iso10T. The average nucleotide identity and digital
DNA-DNA hybridization value between strains 4H37-19T and
3HC-13 and members within genus Corynebacterium were
all below 95% and 70%, respectively. The genomic G + C content
of strains 4H37-19T and 3HC-13 was 52.5%. Diphosphatidylglycerol
(DPG), phosphatidylglycerol (PG), phosphatidylinositol
(PI), phosphatidylcholine, and phosphatidyl inositol
mannosides (PIM) were the major polar lipids, with
C18:1ω9c, C16:0, and C18:0 as the major fatty acids, and MK-8
(H4), MK-8(H2), and MK-9(H2) as the predominant respiratory
quinones. The major whole cell sugar was arabinose,
and the cell wall included mycolic acids. The cell wall peptidoglycan
contained meso-diaminopimelic acid (meso-DAP).
The polyphasic taxonomic data shows that these two strains
represent a novel species of the genus Corynebacterium, for
which the name Corynebacterium poyangense sp. nov. is
proposed. The type strain of Corynebacterium poyangense
is 4H37-19T (=GDMCC 1.1738T = KACC 21671T).
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Valid and accepted novel bacterial taxa isolated from non-domestic animals and taxonomic revisions published in 2023 Erik Munson, Claire R. Burbick, Sara D. Lawhon, Trinity Krueger, Elena Ruiz-Reyes, Romney M. Humphries Journal of Clinical Microbiology.2024;[Epub] CrossRef
Validation List no. 212. Valid publication of new names and new combinations effectively published outside the IJSEM Aharon Oren, Markus Göker
International Journal of Systematic and Evolutionary Microbiology
.2023;[Epub] CrossRef
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Two Gram-stain-positive, oxidase-negative, catalase-positive,
and coccus-shaped bacterial strains, designated CBA3104T
and CBA3105T, were isolated from kimchi. Strain CBA3104T
and CBA3105T grew at 10–35°C (optimum, 25°C and 30°C,
respectively), at pH 6.0–8.5 (optimum, pH 6.5), and in the
presence of 0–15% (w/v) NaCl (optimum, 5%). A phylogenetic
analysis based on 16S rRNA gene sequences revealed
that strain CBA3104T formed a distinct phylogenetic lineage
within the genus Brachybacterium whereas strain CBA3105T
was closely positioned with Brachybacterium halotolerans
MASK1Z-5T. The 16S rRNA gene sequence similarity between
strains CBA3104T and CBA3105T was 99.9%, but ANI
and dDDH values between strains CBA3104T and CBA3105T
were 93.61% and 51.5%, respectively. Strain CBA3104T showed
lower ANI and dDDH values than species delineation against
three closely related strains and type species of the genus
Brachybacterium, however, strain CBA3105T showed 96.63%
ANI value and 69.6% dDDH value with Brachybacterium
halotolerans MASK1Z-5T. Among biochemical analysis results,
strain CBA3104T could uniquely utilize bromo-succinic
acid whereas only strain CBA3105T was positive for alkaline
phosphatase and α-fucosidase among two novel strains, closely
related strains, and type species of the genus Brachybacterium.
Compared with strain CBA3105T and Brachybacterium
halotolerans JCM 34339T, strain CBA3105T was differentially
positive for acid production of D-arabinose, D-adonitol, and
potassium 5-ketogluconate and enzyme activity of β-glucuronidase.
Both strains contained menaquinone-7 as the dominant
quinone. The cell-wall peptidoglycan of two novel
strains contained meso-diaminopimelic acid. The major fatty
acids of strains CBA3104T and CBA3105T were anteiso-C15:0,
anteiso-C17:0, and iso-C16:0. The major polar lipids of both
strains were phosphatidylglycerol and diphosphatidylglycerol.
Strain CBA3104T possessed a uniquely higher abundance of
tRNA (97 tRNAs) than four Brachybacterium strains used for
comparative taxonomic analysis (54–62 tRNAs). Both the
CBA3104T and CBA3105T strain harbored various oxidoreductase,
transferase, hydrolase, and lyase as strain-specific
functional genes compared to closely related strains and Brachybacterium
type species. The results of biochemical/physiological,
chemotaxonomic, and genomic analyses demonstrated
that strains CBA3104T and CBA3105T represent a novel species
of the genus Brachybacterium and a novel subspecies of
B. halotolerans, respectively, for which the names Brachybacterium
kimchii sp. nov. and B. halotolerans subsp. kimchii
subsp. nov. are proposed. The type strains of the novel
species and the novel subspecies are CBA3104T (= KCCM
43417T = JCM 34759T) and CBA3105T (= KCCM 43418T =
JCM 34760T), respectively.
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Metagenomic Insights into the Taxonomic and Functional Features of Traditional Fermented Milk Products from Russia Alexander G. Elcheninov, Kseniya S. Zayulina, Alexandra A. Klyukina, Mariia K. Kremneva, Ilya V. Kublanov, Tatiana V. Kochetkova Microorganisms.2023; 12(1): 16. CrossRef
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International Journal of Systematic and Evolutionary Microbiology
.2022;[Epub] CrossRef
Complete Genome Sequence of
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Shiga toxin-producing Escherichia coli (STEC) is a foodborne
pathogen capable of causing illness in humans. In a previous
study, our group showed that a STEC isolate belonging to
O22:H8 serotype (strain 154) can interfere with STEC O157:H7
colonization both in vitro and in vivo. Using whole-genome
sequencing and genomic comparative, we predicted a subset
of genes acquired by O22:H8 strain 154 through horizontal
gene transfer that might be responsible for the phenotype
previously described by our group. Among them were identified
genes related to the pathogenesis of non-LEE (locus of
enterocyte effacement) STEC, specific metabolic processes,
antibiotic resistance and genes encoding for the T6SS-1 that
is related to inter-bacterial competition. In addition, we showed
that this strain carries stx1c and stx2dact, a mucus-inducible
variant. The results obtained in this study provide insights
into STEC genomic plasticity and the importance of genomic
islands in the adaptation and pathogenesis of this
pathogen.
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Ubiquitin is highly conserved in most eukaryotes and involved
in diverse physiological processes, including cell division, protein
quality control, and protein degradation mediated by the
ubiquitin-proteasome system after heat shock, glucose-starvation,
and oxidative stress. However, the role of the ubiquitin
gene UBI4, which contains five consecutive head-to-tail ubiquitin
repeats, in meiosis has not been investigated. In this
study, we show that the Saccharomyces cerevisiae polyubiquitin
precursor gene, UBI4, is required to promote synaptonemal
complex (SC) formation and suppress excess doublestrand
break formation. Moreover, the proportion of Zip1
polycomplexes, which indicate abnormal SC formation, in
cells with a mutation in UBI4 (i.e., ubi4Δ cells) is higher than
that of wild-type cells, implying that the UBI4 plays an important
role in the early meiotic prophase I. Interestingly, although
ubi4Δ cells rarely form full-length SCs in the pachytene
stage of prophase I, the Zip3 foci are still seen, as in
wild-type cells. Moreover, ubi4Δ cells proficiently form crossover
and noncrossover products with a slight delay compared
to wild-type cells, suggesting that UBI4 is dispensable in SCcoupled
recombination. Our findings demonstrate that UBI4
exhibits dual functions that are associated with both positive
and negative roles in SC formation and recombination during
meiosis.
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The deubiquitinase Usp7 in
Drosophila melanogaster
is required for synaptonemal complex maintenance
Cathleen M. Lake, Jennifer Gardner, Salam Briggs, Zulin Yu, Grace McKown, R. Scott Hawley Proceedings of the National Academy of Sciences.2024;[Epub] CrossRef
Nitric oxide (NO) is a reactive nitrogen species (RNS) that
plays a vital role in regulating inflammatory processes. Under
abnormal conditions, excessive NO levels can promote the
oxidation of cellular components, which may cause or exacerbate
diseases such as hypertension, cardiovascular dysfunction,
and inflammatory bowel disease (IBD). Previous
studies have shown that reducing NO levels in the lumen can
attenuate the clinical symptoms of IBD. Thus, we aimed to
identify bacteria that can reduce RNS and that can be used
as valuable probiotics. In this study, we isolated bacteria resistant
to nitrite stress from human feces and used 16S and
whole-genome sequencing to identify them as Lactiplantibacillus
plantarum LP7 (LP7). The ability to survive at high
nitrite levels and to decrease them was greater in the LP7 strain
than in the reference strain L. plantarum ATCC14917 (ATCC-
14917). To characterize the LP7 genome in more detail, we
performed a comparative genome analysis. However, the unique
genes that directly confer the ability to withstand nitrite
stress were not present in the LP7 genome. Furthermore, we
performed transcriptomic analysis of LP7 and ATCC14917
cells treated with nitrite. We found that the expression levels
of genes involved in the cell division process were induced in
LP7, which showed a more regular rod-shape than ATCC-
14917. This could explain why LP7 can survive better than
ATCC14917 under nitrite stress. Based on its ability to survive
better in nitrite stress and decrease nitrite concentration,
we suggest that LP7 could be a valuable probiotic strain.
Three major proteases, elastase B (LasB), protease IV (PIV),
and elastase A (LasA) expressed in Pseudomonas aeruginosa
play important roles in infections and pathogeneses. These
are activated by a proteolytic cascade initiated by the activation
of LasB. In this study, we investigated whether LasB
could be inhibited using its propeptide (LasBpp). Although
LasA and PIV were inhibited by their propeptides, LasB was
not inhibited by purified LasBpp because LasB degraded LasBpp.
To address this problem, mutant LasBpp variants were constructed
to obtain a mutant LasBpp resistant to LasB degradation.
A C-terminal deletion series of LasBpp was tested in
vivo, and two positive candidates, T2 and T2-1, were selected.
However, both caused growth retardation and were unstably
expressed in vivo. Since deleting the C-terminal end of LasBpp
significantly affected its stable expression, substitution mutations
were introduced at the two amino acids near the
truncation site of T2-1. The resulting mutants, LasBppE172D,
LasBppG173A, and LasBppE172DG173A, significantly diminished LasB
activity when overexpressed in vivo and were stably expressed
in MW1, a quorum sensing mutant that does not produce
LasB. In vitro analysis showed that purified LasBppE172DG173A
inhibited LasB activity to a small extent. Summarizing, Cterminal
modification of LasBpp profoundly affected the stable
expression of LasBpp, and little enhanced the ability of
LasBpp to resist degradation by LasB.
Metabolic abnormalities are one of the main hallmarks of
cancer and are associated with chemoresistance. Therefore,
targeting the metabolic reprogramming of cancer cells has
the potential to overcome chemoresistance. Probiotic-derived
extracellular vesicles (EVs) play important roles in biological
function and intracellular communication. However, the inhibitory
effect of Lactobacillus plantarum-derived EVs (LpEVs)
on colorectal cancer (CRC) cells has not yet been elucidated.
This study clearly revealed that increased glycolysis in 5-fluorouracil
(5-FU)-resistant CRC cells (CRC/5FUR) is directly
related to chemoresistance and that the metabolic shift reversed
by LpEVs inhibits cancer cell proliferation and eventually
leads to apoptosis. Pyruvate dehydrogenase kinase 2
(PDK2), one of the crucial enzymes for enhancing glycolysis,
was upregulated in CRC/5FUR cells. In our study, LpEVs sensitized
CRC/5FUR cells to 5-FU by attenuating PDK2 expression
in p53-p21-dependent metabolic signaling, thereby
circumventing 5-FU resistance. We demonstrated the effect
of cellular responses to 5-FU by modifying the PDK2
expression level in both 5-FU-sensitive parental CRC and 5-
FU resistant CRC cell lines. Finally, we revealed that the PDK2
signaling pathway can potentially be targeted using LpEVs
treatment to overcome chemoresistant CRC, thereby providing
a potential strategy for CRC treatment by intervening in
tumor metabolism.
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Bacteriophages employ diverse mechanisms to facilitate the
proliferation of bacteriophages. The Salmonella-infecting
phage SPN3US contains a putative N-acetyltransferase, which
is widely found in bacteriophages. However, due to low sequence
similarity to the N-acetyltransferases from bacteria
and eukaryotic cells, the structure and function of phage-encoded
acetyltransferases are mainly unknown. This study
determines the crystal structure of the putative N-acetyltransferase
of SPN3US in complex with acetyl-CoA. The crystal
structure showed a novel homodimeric arrangement stabilized
by exchanging the C-terminal α-helix within the dimer.
The following biochemical analyses suggested that the phageencoded
acetyltransferase might have a very narrow substrate
specificity. Further studies are required to reveal the biochemical
activity, which would help elucidate the interaction
between the phage and host bacteria in controlling pathogenic
bacteria.
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Bacteria exist in natural environments for most of their life as
complex, heterogeneous, and multicellular aggregates. Under
these circumstances, critical cell functions are controlled by
several signaling molecules known as quorum sensing (QS)
molecules. In Gram-positive bacteria, peptides are deployed
as QS molecules. The development of antibodies against such
QS molecules has been identified as a promising therapeutic
intervention for bacterial control. Hence, the identification of
QS peptides has received considerable attention. Availability
of a fast and reliable predictive model to effectively identify QS
peptides can help the existing high throughput experiments.
In this study, a stacked generalization ensemble model with
Gradient Boosting Machine (GBM)-based feature selection,
namely EnsembleQS was developed to predict QS peptides
with high accuracy. On selected GBM features (791D), the
EnsembleQS outperformed finely tuned baseline classifiers
and demonstrated robust performance, indicating the superiority
of the model. The accuracy of EnsembleQS is 4% higher
than those resulting from ensemble model on hybrid dataset.
When evaluating an independent data set of 40 QS peptides,
the EnsembleQS model showed an accuracy of 93.4% with
Matthew’s Correlation Coefficient (MCC) and area under the
ROC curve (AUC) values of 0.91 and 0.951, respectively. These results suggest that EnsembleQS will be a useful computational
framework for predicting QS peptides and will efficiently
support proteomics research. The source code and all
datasets used in this study are publicly available at https://
github.com/proteinexplorers/EnsembleQS.
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