Koketso Desiree Mazwi, Kgaugelo Edward Lekota, Barbara Akofo Glover, Francis Babaman Kolo, Ayesha Hassim, Jenny Rossouw, Annelize Jonker, Justnya Maria Wojno, Giuseppe Profiti, Pier Luigi Martelli, Rita Casadio, Katiuscia Zilli, Anna Janowicz, Francesca Marotta, Giuliano Garofolo, Henriette van Heerden
J. Microbiol. 2024;62(9):759-773. Published online July 22, 2024
Brucellosis is an economically important zoonotic disease affecting humans, livestock, and wildlife health globally and especially in Africa. Brucella abortus and B. melitensis have been isolated from human, livestock (cattle and goat), and wildlife (sable) in South Africa (SA) but with little knowledge of the population genomic structure of this pathogen in SA. As whole genome sequencing can assist to differentiate and trace the origin of outbreaks of Brucella spp.
strains, the whole genomes of retrospective isolates (n = 19) from previous studies were sequenced. Sequences were analysed using average nucleotide identity (ANI), pangenomics, and whole genome single nucleotide polymorphism (wgSNP) to trace the geographical origin of cases of brucellosis circulating in human, cattle, goats, and sable from different provinces in SA. Pangenomics analysis of B. melitensis (n = 69) and B. abortus (n = 56) was conducted with 19 strains that included B. abortus from cattle (n = 3) and B. melitensis from a human (n = 1), cattle (n = 1), goat (n = 1), Rev1 vaccine strain (n = 1), and sable (n = 12).
Pangenomics analysis of B. melitensis genomes, highlighted shared genes, that include 10 hypothetical proteins and genes that encodes for acetyl-coenzyme A synthetase (acs), and acylamidase (aam) amongst the sable genomes. The wgSNP analysis confirmed the B. melitensis isolated from human was more closely related to the goat from the Western Cape Province from the same outbreak than the B.
melitensis cattle sample from different cases in the Gauteng Province. The B.
melitensis sable strains could be distinguished from the African lineage, constituting their own African sub-clade. The sequenced B. abortus strains clustered in the C2 lineage that is closely related to the isolates from Mozambique and Zimbabwe. This study identified genetically diverse Brucella spp.
among various hosts in SA. This study expands the limited known knowledge regarding the presence of B. melitensis in livestock and humans in SA, further building a foundation for future research on the distribution of the Brucella spp. worldwide and its evolutionary background.
Adenovirus (Ad) is a ubiquitous pathogen capable of infecting a wide range of animals and humans. Human Adenovirus (HAdV) can cause severe infection, particularly in individuals with compromised immune systems. To date, over 110 types of HAdV have been classified into seven species from A to G, with the majority belonging to the human adenovirus species D (HAdV-D). In the HAdV-D, the most significant factor for the creation of new adenovirus types is homologous recombination between viral genes involved in determining the virus tropism or evading immune system of host cells. The E4 gene, consisting of seven Open Reading Frames (ORFs), plays a role in both the regulation of host cell metabolism and the replication of viral genes. Despite long-term studies, the function of each ORF remains unclear. Based on our updated information, ORF2, ORF3, and ORF4 have been identified as regions with relatively high mutations compared to other ORFs in the E4 gene, through the use of in silico comparative analysis. Additionally, we managed to visualize high mutation sections, previously undetectable at the DNA level, through a powerful amino acid sequence analysis tool known as proteotyping. Our research has revealed the involvement of the E4 gene in the evolution of human adenovirus, and has established accurate sequence information of the E4 gene, laying the groundwork for further research.
Human adenoviruses (HAdVs) can infect various epithelial mucosal cells, ultimately causing different symptoms in infected organ systems. With more than 110 types classified into seven species (A-G), HAdV-D species possess the highest number of viruses and are the fastest proliferating. The emergence of new adenovirus types and increased diversity are driven by homologous recombination (HR) between viral genes, primarily in structural elements such as the penton base, hexon and fiber proteins, and the E1 and E3 regions. A comprehensive analysis of the HAdV genome provides valuable insights into the evolution of human adenoviruses and identifies genes that display high variation across the entire genome to determine recombination patterns. Hypervariable regions within genetic sequences correlate with functional characteristics, thus allowing for adaptation to new environments and hosts. Proteotyping of newly emerging and already established adenoviruses allows for prediction of the characteristics of novel viruses. HAdV-D species evolved in a direction that increased diversity through gene recombination. Bioinformatics analysis across the genome, particularly in highly variable regions, allows for the verification or re-evaluation of recombination patterns in both newly introduced and pre-existing viruses, ultimately aiding in tracing various biological traits such as virus tropism and pathogenesis. Our research does not only assist in predicting the emergence of new adenoviruses but also offers critical guidance in regard to identifying potential regulatory factors of homologous recombination hotspots.
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In Silico Intensive Analysis for the E4 Gene Evolution of Human Adenovirus Species D Chanhee Lee, Anyeseu Park, Jeong Yoon Lee Journal of Microbiology.2024; 62(5): 409. CrossRef
It was reported that LAMMER kinase in Schizosaccharomyces pombe plays an important role in cation-dependent and
galactose-specific flocculation. Analogous to other flocculating yeasts, when cell wall extracts of the Δlkh1 strain were treated
to the wild-type strain, it displayed flocculation. Gas2, a 1,3-β-glucanosyl transferase, was isolated from the EDTA-extracted
cell-surface proteins in the Δlkh1 strain. While disruption of the gas2+ gene was not lethal and reduced the flocculation
activity of the Δlkh1 strain, the expression of a secreted form of Gas2, in which the GPI anchor addition sequences had been
removed, conferred the ability to flocculate upon the WT strain. The Gas2-mediated flocculation was strongly inhibited by
galactose but not by glucose. Immunostaining analysis showed that the cell surface localization of Gas2 was crucial for the
flocculation of fission yeast. In addition, we identified the regulation of mbx2+ expression by Lkh1 using RT-qPCR. Taken
together, we found that Lkh1 induces asexual flocculation by regulating not only the localization of Gas2 but also the transcription
of gas2+ through Mbx2.
The consumption of fresh produce has led to increase in antibiotic-resistant (AR) Salmonella outbreaks. In this study, indigenous
Salmonella was isolated from a total of two hundred-two samples including fresh produce and agricultural environmental
samples in Korea. After biochemical confirmation using the Indole, Methyl Red, Voges-Proskauer, Citrate tests, presumable
Salmonella isolates were identified by 16S rRNA sequencing. Identified Salmonella isolates were evaluated for antibiotic
susceptibility against twenty-two antibiotics. The specificity and the efficiency of plating (EOP) of vB_SalS_KFSSM were
evaluated against fifty-three bacterial strains. Twenty-five suspected Salmonella were isolated and confirmed by the positive
result for methyl red and citrate, of which ten were identified as Salmonella spp. through 16S rRNA gene sequencing. Eight
Salmonella isolates (4.0%, n = 8/202) were resistant to at least one antibiotic, among which five were multi-drug resistant. As
a lytic phage against Salmonella spp. CMGS-1, vB_SalS_KFSSM was isolated from cow manure. The phage was observed as
a tailed phage belonging to the class Caudoviricetes. It exhibited an intra-broad specificity against four indigenous AR Salmonella
isolates, two indigenous Salmonella isolates, and five other Salmonella serotypes with great efficiencies (EOP ≥ 0.75).
Thus, this study suggested the potential of vB_SalS_KFSSM to combat indigenous AR Salmonella.
The global public health burden of bacterial antimicrobial resistance (AMR) is intensified by Gram-negative bacteria,
which have an additional membrane, the outer membrane (OM), outside of the peptidoglycan (PG) cell wall. Bacterial twocomponent
systems (TCSs) aid in maintaining envelope integrity through a phosphorylation cascade by controlling gene
expression through sensor kinases and response regulators. In Escherichia coli, the major TCSs defending cells from envelope
stress and adaptation are Rcs and Cpx, which are aided by OM lipoproteins RcsF and NlpE as sensors, respectively. In
this review, we focus on these two OM sensors. β-Barrel assembly machinery (BAM) inserts transmembrane OM proteins
(OMPs) into the OM. BAM co-assembles RcsF, the Rcs sensor, with OMPs, forming the RcsF-OMP complex. Researchers
have presented two models for stress sensing in the Rcs pathway. The first model suggests that LPS perturbation stress
disassembles the RcsF-OMP complex, freeing RcsF to activate Rcs. The second model proposes that BAM cannot assemble
RcsF into OMPs when the OM or PG is under specific stresses, and thus, the unassembled RcsF activates Rcs. These two
models may not be mutually exclusive. Here, we evaluate these two models critically in order to elucidate the stress sensing
mechanism. NlpE, the Cpx sensor, has an N-terminal (NTD) and a C-terminal domain (CTD). A defect in lipoprotein trafficking
results in NlpE retention in the inner membrane, provoking the Cpx response. Signaling requires the NlpE NTD, but
not the NlpE CTD; however, OM-anchored NlpE senses adherence to a hydrophobic surface, with the NlpE CTD playing
a key role in this function.
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Pseudomonas savastanoi pv. glycinea (Psg, also named P.
syringae pv. glycinea and P. amygdali pv. glycinea) is the
causative agent of bacterial blight in soybean. The identification
of virulence factors is essential for understanding
the pathogenesis of Psg. In this study, a mini-Tn5 transposon
mutant library of Psg strain PsgNC12 was screened on soybean,
and one low-virulent mini-Tn5 mutant, designated as
4573, was identified. Sequence analysis of the 4573-mutant
revealed that the mini-Tn5 transposon was inserted in the
Psg_2795 gene. Psg_2795 encodes a FimC-domain protein
that is highly conserved in Pseudomonas. Further analysis
revealed that the mutation and knockout of Psg_2795 results
in a reduced virulence phenotype on soybean, decreased motility,
weakened bacterial attachment to a glass surface and
delayed the population growth within soybean leaves. The
phenotype of the 4573-mutant could be complemented nearly
to wild-type levels using an intact Psg_2795 gene. Collectively,
our results demonstrate that Psg_2795 plays an important
role in the virulence, motility, attachment and the population
growth of PsgNC12 in soybean. This finding provides a new
insight into the function of periplasmic chaperone proteins
in a type I pilus and provides reference information for identifying
Psg_2795 homologues in P. savastanoi and other
bacteria.
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Genetic variation in eukaryotes is mediated during meiosis by
the exchange of genetic material between homologous chromosomes
to produce recombinant chromosomes. Cohesin is
essential to promote proper chromosome segregation, chromosome
morphogenesis, and recombination in meiotic cells.
Cohesin consists of three main subunits–Smc1, Smc3, and the
kleisin subunit Mcd1/Scc1 (Rec8 in meiosis)–and cohesin accessory
factors. In Saccharomyces cerevisiae, the cohesin regulatory
subunit Pds5 plays a role in homolog pairing, meiotic
axis formation, and interhomolog recombination. In this
study, we examine the prophase functions of Pds5 by performing
physical analysis of recombination and three-dimensional
high-resolution microscopy analysis to identify its roles in
meiosis-specific recombination and chromosome morphogenesis.
To investigate whether Pds5 plays a role in mitoticlike
recombination, we inhibited Mek1 kinase activity, which result ed in switching to sister template bias by Rad51-dependent
recombination. Reductions in double-strand breaks
and crossover products and defective interhomolog recombination
occurred in the absence of Pds5. Furthermore, recombination
intermediates, including single-end invasion
and double-Holliday junction, were reduced in the absence
of Pds5 with Mek1 kinase inactivation compared to Mek1
kinase inactivation cells. Interestingly, the absence of Pds5 result ed in increasing numbers of chromosomes with hypercompaction
of the chromosome axis. Thus, we suggest that
Pds5 plays an essential role in recombination by suppressing
the pairing of sister chromatids and abnormal compaction
of the chromosome axis.
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RPA interacts with Rad52 to promote meiotic crossover and noncrossover recombination Jeong H Joo, Soogil Hong, Mika T Higashide, Eui-Hwan Choi, Seobin Yoon, Min-Su Lee, Hyun Ah Kang, Akira Shinohara, Nancy Kleckner, Keun P Kim Nucleic Acids Research.2024; 52(7): 3794. CrossRef
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Middle East Respiratory Syndrome coronavirus (MERS-CoV),
a contagious zoonotic virus, causes severe respiratory infection
with a case fatality rate of approximately 35% in humans.
Intermittent sporadic cases in communities and healthcare
facility outbreaks have continued to occur since its first identification
in 2012. The World Health Organization has declared
MERS-CoV a priority pathogen for worldwide research
and vaccine development due to its epidemic potential and
the insufficient countermeasures available. The Coalition for
Epidemic Preparedness Innovations is supporting vaccine development
against emerging diseases, including MERS-CoV,
based on platform technologies using DNA, mRNA, viral vector,
and protein subunit vaccines. In this paper, we review the
usefulness and structure of a spike glycoprotein as a MERSCoV
vaccine candidate molecule, and provide an update on
the status of MERS-CoV vaccine development. Vaccine candidates
based on both DNA and viral vectors coding MERSCoV
spike gene have completed early phase clinical trials. A
harmonized approach is required to assess the immunogenicity
of various candidate vaccine platforms. Platform technologies
accelerated COVID-19 vaccine development and can
also be applied to developing vaccines against other emerging
viral diseases.
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Eukaryotic genomes contain many duplicated genes closely
located with each other, such as the hexose transporter (HXT)
genes in Saccharomyces cerevisiae. They can potentially recombine
via single-strand annealing (SSA) pathway. SSA between
highly divergent sequences generates heteroduplex
DNA intermediates with many mismatches, which can be
corrected by mismatch repair (MMR), resulting in recombinant
sequences with a single junction point. In this report,
we demonstrate that SSA between HXT1 and HXT4 genes
in MMR-deficient yeast cells produces recombinant genes
with multiple-junctions resulting from alternating HXT1 and
HXT4 tracts. The mutations in MMR genes had differential
effects on SSA frequencies; msh6Δ mutation significantly
stimulated SSA events, whereas msh2Δ and msh3Δ slightly
suppressed it. We set up an assay that can identify a pair of
recombinant genes derived from a single heteroduplex DNA.
As a result, the recombinant genes with multiple-junctions
were found to accompany genes with single-junctions. Based
on the results presented here, a model was proposed to generate
multiple-junctions in SSA pathway involving an alternative
short-patch repair system.
Brucella, the bacterial agent of common zoonotic brucellosis,
primarily infects specific animal species. The Brucella outer
membrane proteins (Omps) are particularly attractive for developing
vaccine and improving diagnostic tests and are associated
with the virulence of smooth Brucella strains. Omp16
is a homologue to peptidoglycan-associated lipoproteins (Pals),
and an omp16 mutant has not been generated in any Brucella
strain until now. Very little is known about the functions and
pathogenic mechanisms of Omp16 in Brucella. Here, we confirmed
that Omp16 has a conserved Pal domain and is highly
conserved in Brucella. We attempted to delete omp16 in Brucella
suis vaccine strain 2 (B. suis S2) without success, which
shows that Omp16 is vital for Brucella survival. We acquired
a B. suis S2 Omp16 mutant via conditional complementation.
Omp16 deficiency impaired Brucella outer membrane integrity
and activity in vitro. Moreover, inactivation of Omp16
decreased bacterial intracellular survival in macrophage
RAW 264.7 cells. B. suis S2 and its derivatives induced marked
expression of IL-1β, IL-6, and TNF-α mRNA in Raw 264.7
cells. Whereas inactivation of Omp16 in Brucella enhanced
IL-1β and IL-6 expression in Raw 264.7 cells. Altogether, these
findings show that the Brucella Omp16 mutant was obtained
via conditional complementation and confirmed that Omp16
can maintain outer membrane integrity and be involved in
bacterial virulence in Brucella in vitro and in vivo. These results
will be important in uncovering the pathogenic mechanisms
of Brucella.
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Pal Affects the Proliferation in Macrophages and Virulence of Brucella, and as Mucosal Adjuvants, Provides an Effective Protection to Mice Against Salmonella Enteritidis Yubin Chen, Yanfang Fu, Lingcong Kong, Fengjie Wang, Xiaowei Peng, Zhiqiang Zhang, Qiumei Shi, Qingmin Wu, Tonglei Wu Current Microbiology.2023;[Epub] CrossRef
Clearance of bacteria from lymph nodes in sheep immunized with Brucella suis S2 vaccine is associated with M1 macrophage activation Si Chen, Yuanyuan Chen, Zizhuo Jiao, Chengqiang Wang, Dantong Zhao, Yongbin Liu, Wenguang Zhang, Shihua Zhao, Bin Yang, Qinan Zhao, Shaoyin Fu, Xiaolong He, Qiaoling Chen, Churiga Man, Guoying Liu, Xuefeng Wei, Li Du, Fengyang Wang Veterinary Research.2023;[Epub] CrossRef
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Enterovirus A71 (EV71), the main etiological agent of handfoot-
mouth disease (HFMD), circulates in many areas of the
world and has caused large epidemics since 1997, especially
in the Asia-Pacific region. In this study, we determined the
full-genome sequence of CMC718, a newly isolated EV71
strain in Korea. The CMC718 genome was 7,415 nucleotides
in length and was confirmed by whole-genome phylogenetic
analysis to belong to the B5 genotype. In particular, CMC718
demonstrated maximum identity with strain M988 of the B5
genotype and numerous amino acid variants were detected
in the 3D domain of the viral protein P3, which is consistent
with the mutation pattern of a B5 strain isolated in 2012–2013.
Comparison of the CMC718 sequence with other EV71 reference
strains confirmed the relationship and genetic variation
of CMC718. Our study was a full-genome sequence analysis
of the first EV71 strain of the B5 genotype isolated in
South Korea. This information will be a valuable reference
for the development of methods for the detection of recombinant
viruses, the tracking of infections, and the diagnosis
of EV71.
Byeong Seob Oh , Ji-Sun Kim , Seung Yeob Yu , Seoung Woo Ryu , Seung-Hwan Park , Se Won Kang , Jam-Eon Park , Seung-Hyeon Choi , Kook-Il Han , Keun Chul Lee , Mi Kyung Eom , Min Kuk Suh , Han Sol Kim , Dong Ho Lee , Hyuk Yoon , Byung-Yong Kim , Je Hee Lee , Jung-Sook Lee , Ju Huck Lee
J. Microbiol. 2020;58(2):99-104. Published online January 29, 2020
An obligately anaerobic, Gram-stain-negative, non-motile,
non-spore-forming, and coccobacilli-shaped bacterial strain,
designated KGMB03119T, was isolated from human faeces
from a Korean. Phylogenetic analysis based on the 16S rRNA
gene sequence revealed that the isolate was a member of the
genus Sutterella and most closely related to Sutterlla wadsworthensis
KCTC 15691T (96.8% 16S rRNA gene sequence
similarity). The DNA G + C content of strain KGMB03119T
was 58.3 mol% as determined from its whole genome sequence.
Strain KGMB03119T was asaccharolytic, catalase-positive,
oxidase- and urease-negative. Furthermore, the isolate
was positive for alkaline phosphatase, leucine arylamidase,
acid phosphatase, arginine arylamidase, alanine arylamidase,
and glycine arylamidase. The major cellular fatty acids (> 10%)
of the isolate were C18:1ω9c and C16:0. Methylmenaquinone-5
(MMK-5, 100%) was the predominant isoprenoid quinone
in the isolate. Based on the phylogenetic, physiological, and
chemotaxonomic characteristics, strain KGMB03119T represents
a novel species, for which the name Sutterella faecalis
sp. nov. is proposed. The type strain is KGMB03119T (= KCTC
15823T = NBRC 114254T).
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The Chikungunya virus (CHIKV) belongs to the Alphavirus
genus of Togaviridae family and contains a positive-sense
single stranded RNA genome. Infection by this virus mainly
causes sudden high fever, rashes, headache, and severe joint
pain that can last for several months or years. CHIKV, a mosquito-
borne arbovirus, is considered a re-emerging pathogen
that has become one of the most pressing global health
concerns due to a rapid increase in epidemics. Because handling
of CHIKV is restricted to Biosafety Level 3 (BSL-3) facilities,
the evaluation of prophylactic vaccines or antivirals
has been substantially hampered. In this study, we first identified
the whole structural polyprotein sequence of a CHIKV
strain isolated in South Korea (KNIH/2009/77). Phylogenetic
analysis showed that this sequence clustered within the East/
Central/South African CHIKV genotype. Using this sequence
information, we constructed a CHIKV-pseudotyped lentivirus
expressing the structural polyprotein of the Korean
CHIKV isolate (CHIKVpseudo) and dual reporter genes of
green fluorescence protein and luciferase. We then developed
a pseudovirus-based neutralization assay (PBNA) using
CHIKVpseudo. Results from this assay compared to those
from the conventional plaque reduction neutralization test
showed that our PBNA was a reliable and rapid method to
evaluate the efficacy of neutralizing antibodies. More importantly,
the neutralizing activities of human sera from CHIKVinfected
individuals were quantitated by PBNA using CHIKVpseudo.
Taken together, these results suggest that our PBNA
for CHIKV may serve as a useful and safe method for testing
the neutralizing activity of antibodies against CHIKV
in BSL-2 facilities.
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A strictly anaerobic bacterium, designated as strain KGMB-
03357T, was isolated from the faeces of a healthy Korean selected
by Bundang Seoul National University based on health
status. Cells of strain KGMB03357T are Gram-stain-positive,
non-motile, non-spore-forming, and observed as straight or
curved rods. The isolate grew at 10–45°C (optimum temperature
of 40°C) and a pH range of 5.1–10.5 (optimum pH of
6.8). Analysis of phylogenetic trees based on the 16S rRNA
gene sequences revealed that strain KGMB03357T forms a
lineage within the genus Anaerotignum, and is most closely
related to Anaerotignum lactatifermentans G17T (= KCTC
15066T, 96.1%), Anaerotignum propionicum DSM 1682T (=
KCTC 5582T, 94.9%), Anaerotignum neopropionicum DSM
03847T (= KCTC 15564T, 94.9%), and Anaerotignum aminivorans
SH021T (= KCTC 15705T, 94.8%). The ANI values
between strain KGMB 03357T and members of the genus
Anaerotignum were 73.3–71.0%, which are below the ANI
criterion for interspecies identity. The DNA G + C content
based on the whole-genome sequence is 47.3 mol%. The major
cellular fatty acids of strain KGMB03357T are C16:0, C18:0,
C18:1 cis 9, and anteiso-C15:0. Strain KGMB03357T contains
meso-diaminopimelic acid as the diagnostic amino acid in
the cell wall peptidoglycan. Based on the phenotypic, phylogenetic,
and genomic properties, strain KGMB 03357T represents
a novel species of the genus Anaerotignum, for which
the name Anaerotignum faecicola sp. nov. is proposed. The
type strain is KGMB03357T (= KCTC 15736T = DSM 107953T).
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