First Whole Genome Report of Mangrovibacter phragmitis PSU-3885-11 Isolated from a Patient in Thailand

Abstract

Mangrovibacter phragmitis is a Gram-negative bacterium typically found in plant roots that supports nitrogen fixation in nutrient-poor environments such as mangrove ecosystems. Although primarily found in environmental niches, an unusual case in Thailand of M. phragmitis strain PSU-3885–11 isolated from the sputum of a 29-year-old female patient with spinal tuberculosis. This isolate was initially misidentified as part of the Enterobacter cloacae complex (ECC) by MALDI-TOF. However, WGS subsequently confirmed its correct identity as M. phragmitis. The genome contains 4,651 coding sequences, along with 72 tRNA genes and 1 tmRNA. Moreover, comparative genomic analysis showed 99.32 % average nucleotide identity (ANI) similar to M. phragmitis MP23, and several antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) were identified in the PSU-3885–11 genome which may contribute to its ability to survive in diverse environments, including human hosts. The PSU-3885–11 displayed resistance to beta-lactam antibiotics such as ampicillin and cefotaxime, while remaining sensitive to a wide range of other antibiotics. Key virulence genes including ompA, hcp/tssD, and rpoS, were identified which may play a role in its persistence in human hosts as an opportunistic pathogen. The presence of ribosomally synthesized and post-translationally modified peptides (RiPPs) and bacteriocins indicates the antimicrobial properties that may provide a competitive advantage in both environmental and clinical settings of this strain. Therefore, this study provides valuable insights into the genomic features, antibiotic resistance, and potential pathogenicity of M. phragmitis PSU-3885–11. The findings also emphasize the importance of continued surveillance and genomic analysis of environmental bacteria that may emerge as opportunistic pathogens in human infections.

Full text

First whole genome report of Mangrovibacter phragmitis PSU-3885–11
isolated from a patient in Thailand
Nattarika Chaichana
a
, Thunchanok Yaikhan
a
, Mingkwan Yingkajorn
b
,
Nonthawat Thepsimanon
b
, Sirikan Suwannasin
a
, Kamonnut Singkhamanan
a
,
Sarunyou Chusri
c
, Rattanaruji Pomwised
d
, Monwadee Wonglapsuwan
d
,
Komwit Surachat
a,e,*
a
Department of Biomedical Sciences and Biomedical Engineering, Faculty of Medicine, Prince of Songkla University, Songkhla 90110, Thailand
b
Department of Pathology, Faculty of Medicine, Prince of Songkla University, Songkhla 90110, Thailand
c
Division of Infectious Diseases, Department of Internal Medicine, Faculty of Medicine, Prince of Songkla University, Songkhla 90110, Thailand
d
Division of Biological Science, Faculty of Science, Prince of Songkla University, Songkhla 90110, Thailand
e
Translational Medicine Research Center, Faculty of Medicine, Prince of Songkla University, Songkhla 90110, Thailand
ARTICLE INFO
Keywords:
Whole genome sequencing
Mangrovibacter phragmitis
Draft genome
Bioinformatics
Antibiotic resistance
ABSTRACT
Mangrovibacter phragmitis is a Gram-negative bacterium typically found in plant roots that supports nitrogen
xation in nutrient-poor environments such as mangrove ecosystems. Although primarily found in environ-
mental niches, an unusual case in Thailand of M. phragmitis strain PSU-3885–11 isolated from the sputum of a 29-
year-old female patient with spinal tuberculosis. This isolate was initially misidentied as part of the Enterobacter
cloacae complex (ECC) by MALDI-TOF. However, WGS subsequently conrmed its correct identity as
M. phragmitis. The genome contains 4,651 coding sequences, along with 72 tRNA genes and 1 tmRNA. Moreover,
comparative genomic analysis showed 99.32 % average nucleotide identity (ANI) similar to M. phragmitis MP23,
and several antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) were identied in the PSU-
3885–11 genome which may contribute to its ability to survive in diverse environments, including human hosts.
The PSU-3885–11 displayed resistance to beta-lactam antibiotics such as ampicillin and cefotaxime, while
remaining sensitive to a wide range of other antibiotics. Key virulence genes including ompA, hcp/tssD, and rpoS,
were identied which may play a role in its persistence in human hosts as an opportunistic pathogen. The
presence of ribosomally synthesized and post-translationally modied peptides (RiPPs) and bacteriocins in-
dicates the antimicrobial properties that may provide a competitive advantage in both environmental and
clinical settings of this strain. Therefore, this study provides valuable insights into the genomic features, anti-
biotic resistance, and potential pathogenicity of M. phragmitis PSU-3885–11. The ndings also emphasize the
importance of continued surveillance and genomic analysis of environmental bacteria that may emerge as
opportunistic pathogens in human infections.
1. Introduction
The genus Mangrovibacter is a Gram-negative, rod-shaped, and
facultatively anaerobic bacteria that belongs to the Enterobacteriaceae
family. Mangrovibacter sp. is recognized for its connection to plant roots,
where it typically contributes to enhancing plant growth and health such
as nitrogen xation (Rameshkumar et al., 2010). This process converts
atmospheric nitrogen into a form usable by plants, such as ammonium. It
is especially benecial in nutrient-poor environments, such as mangrove
ecosystems, where it greatly enhances nitrogen availability for host
plants (Alfaro-Espinoza and Ullrich, 2015). Only three known species of
Mangrovibacter were identied and their nucleotides were deposited in
the National Center for Biotechnology Information (NCBI) database
including M. plantisponsor, M. yixingensis, and M. phragmitis. Among
these species, M. plantisponsor was rst identied during a study inves-
tigating plant-microbe interactions in mangrove ecosystems. It was
isolated from the roots of Porteresia coarctata, a wild rice species natu-
rally adapted to saline and waterlogged environments along India’s
* Corresponding author.
E-mail address: komwit.s@psu.ac.th (K. Surachat).
Contents lists available at ScienceDirect
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https://doi.org/10.1016/j.crmicr.2025.100350
Current Research in Microbial Sciences 8 (2025) 100350
Available online 22 January 2025
2666-5174/© 2025 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-
nc-nd/4.0/ ).

coasts. This halophytic plant is renowned for its exceptional salt toler-
ance and ability to thrive in challenging mangrove habitats
(Rameshkumar et al., 2010). Similarly, M. yixingensis was isolated in
2015 from farmland soil in Yixing, Jiangsu Province, China. This species
was identied through 16S rRNA gene sequencing, revealing a close
relationship to the M. plantisponsor (Zhang et al., 2015). In contrast,
M. phragmitis was recovered from the roots of Phragmites karka (tall reed)
in Odisha, India. This bacterium exhibits signicant plant
growth-promoting traits, including nitrogen xation and phosphate
solubilization, making it a promising candidate for agricultural appli-
cations (Behera et al., 2017). Furthermore, its ability to adapt to saline
environments highlights its potential for biotechnological use in man-
aging saline soils. Their genomes also exhibit adaptive features for
survival in specic environments, such as salt tolerance mechanisms in
M. plantisponsor and M. phragmitis (Behera et al., 2017; Rameshkumar
et al., 2010). These species are primarily non-pathogenic and lack sig-
nicant virulence factors, aligning with their roles as benecial plant
symbionts rather than pathogens. Genetic analyses reveal evidence of
horizontal gene transfer, suggesting adaptability to varying environ-
mental conditions. Transmission occurs naturally through soil and
water, facilitating the colonization of host plants. Their plant
growth-promoting traits and environmental adaptability position them
as promising candidates for agricultural applications, including
enhancing crop productivity in nutrient-poor or saline soils, while also
holding potential for broader biotechnological use.
The draft genome sequence of M. phragmitis provides functional
characteristics of oxidative stress, uptake of nutrients, and nitrogen
xation to produce a range of enzymes and secondary metabolites that
may have applications in biotechnology and agriculture (Behera et al.,
2016). Overall, M. phragmitis is typically found in environmental areas
and is rarely associated with human infections. The presence of
M. phragmitis in common is uncommon in humans is uncommon and
could either be an incidental observation or, in rare cases, indicate the
emergence of an opportunistic pathogen, especially in individuals with
underlying health conditions or compromised immune systems. In our
study, this bacterium was unexpectedly isolated from a 29-year-old fe-
male patient presenting with symptoms consistent with spinal tubercu-
losis. Her clinical signs included a gibbous deformity, cold abscess,
paradiscal lesion, anterior vertebral loss, narrowed disc space, and
paravertebral shadows. Moreover, she exhibited tuberculosis-related
symptoms such as loss of appetite, weight loss, and malnutrition, with
a body mass index (BMI) below 18.5, which is a signicant risk factor for
TB infection.
During our one-year surveillance of Enterobacter cloacae complex
(ECC) from patients in Songklanagarind Hospital, we obtained this PSU-
3885–11 strain from the sputum of the patient that was rst mis-
identied as ECC by MALDI-TOF analysis. The whole-genome
sequencing (WGS) is an advanced and powerful tool with accuracy in
current microbiological research that allows for a comprehensive anal-
ysis of the bacterial genome including its gene content, regulatory ele-
ments, and non-coding regions (Quainoo et al., 2017). This approach
offers specic genes responsible for its adaptation to human hosts, po-
tential virulence factors, and antibiotic resistance prole which are
crucial information on the rarity of this strain in clinical settings and
provide valuable insights into its possible role as an opportunistic
pathogen (Sornchuer et al., 2024; Zaghloul and El Halfawy, 2022).
Therefore, our study focuses on the genomic characterization of
M. phragmitis PSU-3885–11 isolated from a patient in Thailand to obtain
a comprehensive understanding of its genetic makeup, potential path-
ogenic traits, and antibiotic resistance prole. The insight information
gained from this study is crucial for enhancing public health strategies of
environmental bacteria, guiding investigations into how these bacteria
may adapt to human hosts and transition into pathogenic roles.
2. Materials and methods
2.1. Patient sample and bacterial isolation
Mangrovibacter phragmitis PSU-3885–11 was isolated from the
sputum of a 29-year-old female patient presenting with symptoms
consistent with spinal tuberculosis at Songklanagarind Hospital. A bone
tissue sample was obtained from the spine and the patient was diag-
nosed with tuberculosis using XpertⓇ MTB/RIF Ultra. The bacterial
isolate was obtained using the BD BACTEC™ MGIT™ culture system.
Bacterial isolate was then identied by All the procedures involving
human participants were carried out following the rules of the Decla-
ration of Helsinki and approved by the Human Research Ethics Com-
mittee (HREC) of Prince of Songkla University (protocol code:
64–284–14–1, date of approval: 9 June 2021).
2.2. Antimicrobial susceptibility testing
Antimicrobial susceptibility testing (AST) of M. phragmitis PSU-
3885–11 was conducted using disk diffusion assay. Antibiotic drugs
were used in this study including amikacin (AK) 10 µg, ampicillin (AM)
10 µg, cefotaxime (CTX) 30 µg, cefoxitin (FOX) 30 µg, ceftazidime (CAZ)
30 µg, ceftriaxone (CRO) 30 µg, cefuroxime (CXM) 30 µg, ciprooxacin
(CIP) 5 µg, trimethoprim-sulfamethoxazole (Co-trimozole; STX) 1.25/
23.75 µg, gentamicin (GM) 10 µg, imipenem (IPM) 10 µg, meropenem
(MEM) 10 µg, piperacillin-tazobactam (TZP) 100/10 µg, ertapenem
(ETP) 10 µg, and chloramphenicol (CL) 30 µg. The results of the AST
were interpreted according to the Clinical & Laboratory Standards
Institute (CLSI) standard (CLSI, 2015).
2.3. Genomic dna extraction and whole-genome sequencing
M. phragmitis PSU-3885–11 was cultured on Luria Bertani (LB) agar
(Himedia, Mumbai, India) and then incubated at 37 ◦C for 24 h A single
colony was then subcultured in Luria Bertani (LB) broth (Himedia,
Mumbai, India) under shaking conditions at 37 ◦C for 4 h. The culture
was centrifuged, and the cell pellet was washed twice with phosphate-
buffered saline (PBS). Genomic DNA (gDNA) was subsequently extrac-
ted from the cell pellet using the ZymoBIOMICS DNA Miniprep Kit
(Zymo Research, Irvine, CA, USA) according to the manufacturer’s
protocol. The quality of the gDNA was assessed using 1 % agarose gel
electrophoresis. The DNA samples were sent to the Beijing Genomics
Institute (BGI) for short-read whole genome sequencing (WGS) using the
MGISEQ-2000 platform.
2.4. Genome assembly, annotation and visualization
For genome assembly, the quality of raw reads was initially assessed
using FASTQC v0.10.0. Subsequently, the reads were quality-ltered
with Trimmomatic v0.32 (Bolger et al., 2014). The high-quality
ltered reads were then de novo assembled using SPAdes v4.0.0
(Bankevich et al., 2012). The annotation was then performed using the
NCBI Prokaryotic Genome Annotation Pipeline (PGAP) (Tatusova et al.,
2016), genome visualization was generated with Proksee v1.0.0a6
(Grant et al., 2023) and functional prediction was conducted using
Rapid Annotations using Subsystems Technology (RAST) (Aziz et al.,
2008).
2.5. Mobile genetic element (MGE) prediction
To predict mobile genetic elements (MGEs) in the M. phragmitis PSU-
3885–11 genome, mobileOG-db was utilized to identify MGEs and their
associated orthologous gene families (Brown et al., 2022).
N. Chaichana et al.
Current Research in Microbial Sciences 8 (2025) 100350
2

2.6. Species identication
To identify the bacterial species, FastANI v1.34 (Jain et al., 2018)
was used to perform a pairwise comparison of M. phragmitis
PSU-3885–11 against the reference genome M. phragmitis MP23
(GCF_001655675.1).
2.7. Sequence analysis
The restriction-modication (R-M) sites were detected using
Restriction-ModicationFinder v1.1 (Roer et al., 2016), applying
thresholds of 95 % minimum identity and 80 % minimum coverage.
Additionally, Ribosomally synthesized and Post translationally modied
Peptides (RiPPs) and bacteriocins were predicted by the Bagel4 server
(van Heel et al., 2018).
2.8. Antimicrobial resistance prole among Mangrovibacter sp.
Antimicrobial resistance genes (ARGs) presented in M. phragmitis
PSU-3885–11 were identied using the Comprehensive Antibiotic
Resistance Database (CARD) with a cutoff of 80 % identity and an E-
value threshold of 1e-5 (McArthur et al., 2013). The
virulence-associated genes were detected by applying an 80 % identity
cutoff and an E-value threshold of 1e-30 against the Virulence Factor
Database (VFDB) (Chen et al., 2012).
3. Results and discussion
3.1. Bacterial identication
The biochemical test results of isolate PSU3885–11 are presented in
Table S1. The results suggest the isolate belongs to the Enterobacter
genus based on its metabolic prole, including positive results for VP,
malonate utilization, gas production, and the fermentation of sucrose,
mannitol, arabinose, and sorbitol. These traits align with typical
Enterobacter characteristics, such as the ability to ferment a wide range
of sugars and produce gas during fermentation (Rogers, 2024). The
negative tests for indole production, H
2
S production, and citrate utili-
zation further support this classication. However, the MALDI-TOF
analysis identied the isolate as Klebsiella oxytoca with a low con-
dence score of 1.7 (Table S1), which is below the standard threshold for
reliable species-level identication (typically ≥2.0) (Panda et al., 2014).
This discrepancy is not unexpected due to the MALDI-TOF relying on
spectral matching with reference databases which may lack sufcient or
accurate entries for rare, novel, or less-studied bacteria (Rychert, 2019).
Hence, high-resolution methods, such as whole genome sequencing
provide greater precision and reliability compared to biochemical tests
and MALDI-TOF. This approach would help clarify whether the isolate
belongs to Enterobacter sp., K. oxytoca, or another closely related species.
Moreover, the use of genomic analysis can offer more precise identi-
cation and provide insights into antimicrobial resistance genes, viru-
lence factors, and metabolic pathways, enabling clinicians to tailor
antibiotic treatment more effectively.
3.2. Antimicrobial susceptibility proles
The antibiotic susceptibility pattern of M. phragmitis PSU-3885–11
against the tested antibiotics is shown in Table 1. The strain PSU-
3885–11 exhibited sensitivity to a range of antibiotics, indicating its
susceptibility to various treatments. However, it showed resistance to
certain antibiotics from Group I, specically beta-lactams including
ampicillin and cefotaxime.
The antibiotic susceptibility results of M. phragmitis PSU-3885–11
reveal a selective resistance prole, particularly within the beta-lactam
group suggesting the presence of resistance mechanisms such as beta-
lactamases that degrade these antibiotics (Mora-Ochomogo and
Lohans, 2021). The resistance to ampicillin and cefoxitin while suscep-
tibility to other antibiotics, highlights the importance of conducting
detailed antibiotic susceptibility testing to guide treatment in clinical
settings.
3.3. Genome features of M. phragmitis PSU-3885–11
The genome of M. phragmitis PSU-3885–11 consists of 5035,050 bp
with a GC content of 50 %. The sequence includes 4651 coding se-
quences (CDS), 72 tRNAs, and 1 tmRNA. The average nucleotide identity
(ANI) of the PSU-3885–11 strain compared to the reference strain M.
phragmitis MP23 (GCF_001655675.1 from the National Center for
Biotechnology Information (NCBI) database) is 99.32 %. ANI analysis,
which compares the genomic similarity between two isolates, is a robust
and widely accepted method for conrming taxonomic relationships.
This result suggested that the limitations of MALDI-TOF could provide
misidentication data that delay appropriate treatment or lead to inef-
fective therapy, potentially exacerbating patient outcomes (Feucherolles
et al., 2019). The comprehensive features of the assembled genome are
presented in Table 2, and the circular representation of the genome is
depicted in Fig. 1A. Notably, a comparative analysis of the PSU-3885–11
genome with four other Mangrovibacter strains including M. phragmitis
MP23, M. yixingensis SaN21–3, M. plantisponsor DSM19579, and Man-
grovibacter sp. MFB 070 from the NCBI database revealed that
PSU-3885–11 shares a common core genome with other Mangrovibacter
strains. It has acquired unique genetic elements, particularly MGEs, that
may enhance its adaptability and potential virulence in human hosts.
Specically, the presence of several transferred regions in the
PSU-3885–11 strain suggests that it has acquired genetic material from
other bacterial species (Fig. 1B). This result suggests a signicant role of
horizontal gene transfer (HGT) in its evolutionary history. These trans-
ferred regions likely contain genes that confer advantageous traits, such
Table 1
Antibiotic susceptibility of Mangrovibacter phragmitis PSU-3885–11.
Group numbers Antibiotics Antibiotic
susceptibility
Group I: Cell wall synthesis
inhibitors
AM (10 µg) R
TZP (100/10 µg) S
CTX (30 µg) S
FOX (30 µg) R
CAZ (30 µg) S
CRO (30 µg) S
CXM (30 µg) S
IPM (10 µg) S
MEM (10 µg) S
ETP (10 µg) S
Group II: Aminoglycosides AK (10 µg) S
GM (10 µg) S
Group III: Fluoroquinolones CIP (5 µg) S
Group IV: Folate synthesis
inhibitors
SXT (1.25/23.75
µg)
S
Group VI: Chloramphenicol CL (30 µg) S
Susceptibility is presented as resistant (R) and sensitive (S).
Table 2
Characteristics of M. phragmitis PSU-3885–11 genome.
Features M. phragmitis PSU-3885–11
Size (bp) 5035,050
Number of Contigs 52
GC Content (%) 50.0
Number of Coding Sequences (CDS) 4651
N50 207,029
L50 7
tRNA 72
tmRNA 1
RAST subsystem 360
ANI value with M. phragmitis MP23 from NCBI (%) 99.32
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Current Research in Microbial Sciences 8 (2025) 100350
3

Fig. 1. Circular genome map of Mangrovibacter phragmitis PSU-3885–11 (A) and comparison of mobile genetic elements (MGEs) in transfer regions (highlighted in
blue) with four other Mangrovibacter strains: M. phragmitis MP23, M. yixingensis SaN21–3, M. plantisponsor DSM 19,579, and Mangrovibacter sp. MFB 070. The ge-
nomes of the comparative strains were obtained from the NCBI database (B). Subsystem coverage of the M. phragmitis PSU-3885–11 genome as annotated by Rapid
Annotations using Subsystems Technology (RAST) (C).
N. Chaichana et al.
Current Research in Microbial Sciences 8 (2025) 100350
4

as antibiotic resistance, virulence factors, or metabolic capabilities that
allow this strain to adapt to new environments, including human hosts
(Hall et al., 2017; Panda et al., 2018).
Moreover, the core subsystem structure of the genome annotated by
Rapid Annotations using Subsystems Technology (RAST) includes genes
associated with carbohydrates (321), protein metabolism (210), viru-
lence, disease, and defense (37), stress response (82), and other func-
tional categories (Fig. 1C). The annotation of M. phragmitis PSU-3885–11
with genes associated with carbohydrate and protein metabolisms re-
veals key metabolic capabilities, indicating its adaptability to diverse
nutrient sources. The annotation of virulence, disease, and defense-
related genes suggests that, although M. phragmitis is primarily an
environmental bacterium, it possesses the genetic arsenal necessary for
pathogenic interactions (Beceiro et al., 2013; Strateva and Mitov, 2011).
This includes genes that may enhance its ability to evade host immune
defenses, adhere to host cells, and cause infections in immunocompro-
mised individuals. These virulence factors, in combination with its
metabolic capabilities, may allow M. phragmitis to transition from an
environmental organism to an opportunistic pathogen in certain con-
ditions. The identication of stress response genes suggests robust sur-
vival mechanisms in hostile environments, including human hosts (Fang
et al., 2016). Additionally, the presence of genes linked to secondary
metabolism, iron acquisition, and phosphate metabolism points to its
capacity for producing secondary metabolites and efciently managing
essential nutrients, which could enhance its competitiveness and path-
ogenicity (Chevrette et al., 2022; Klebba et al., 2021). This compre-
hensive genomic prole underscores the versatility of the strain and its
possible role as an emerging pathogen in immunocompromised
individuals.
3.4. Mobile genetic element (MGE) identication
The MGEs within the genome of M. phragmitis PSU-3885–11 were
identied using the mobileOG-db, a specialized database for categoriz-
ing and understanding mobile genetic components. The distribution and
diversity of these MGEs are depicted in the outermost rings of the cir-
cular genome map shown in Fig. 1A. The result found that the PSU-
3885–11 genome processes various MGE regions including integration
and excision (IE; yellow), transfer (T; light blue), replication, recombi-
nation, and repair (RRR; dark pink), phage (P; light green), and stability,
transfer, or defense (STD; purple). Moreover, the 329 MGEs identied in
M. phragmitis PSU-3885–11 were categorized into the groups illustrated
in Fig. 2. The identication of diverse MGEs further in M. phragmitis PSU-
3885–11 provides signicant insights into its genomic plasticity and
adaptability. The large number of MGEs, particularly RRR (107) and IE
(60), suggests that PSU-3885–11 has a high potential for acquiring
foreign DNA, including antibiotic resistance and virulence genes
through HGT (Jeon et al., 2023; Michaelis and Grohmann, 2023).
Moreover, the signicant presence of phage-related genes also un-
derscores the role of bacteriophages in facilitating genetic diversity and
may contribute to the evolutionary success of the strain (Pfeifer et al.,
2022). The genomic exibility may play a critical role in the ability of
bacteria to adapt in different environments, especially human hosts, and
poses a concern for the horizontal transfer of ARGs to other pathogens in
clinical environments (Emamalipour et al., 2020; Wang et al., 2024).
These ndings suggest the importance of monitoring environmental
bacteria of M. phragmitis PSU-3885–11 for their potential to become
emerging pathogens through the acquisition of novel genetic elements.
3.5. Detection of restriction-modication (R-M) sites
The restriction-modication (RM) systems within the M. phragmitis
PSU-3885–11 genome were thoroughly analyzed. The study identied
three key genes: EcoEI, M.Ecl93I, and M.Ecl884AI, with sequence
identities of 88.29 %, 88.59 %, and 90.97 %, respectively. The EcoEI
gene, encoding a restriction enzyme, is located between 101,254 and
103,695 bp, while the M.Ecl93I and M.Ecl884AI genes, both responsible
for encoding methyltransferases, are situated between 122,784 to
124,290 bp and 99,709 to 101,181 bp, respectively, as detailed in
Table 3. The presence of these R-M systems indicates a mechanism for
defense against foreign DNA, which might provide a selective advantage
by protecting the bacterium from phage infections and other genomic
threats in diverse environments (Rusinov et al., 2018; Vasu and
Nagaraja, 2013).
3.6. Ribosomally synthesized and post translationally modied peptides
(RiPPs) and bacteriocins
Four regions in M. phragmitis PSU-3885–11 were identied as con-
taining RiPPs and bacteriocins, as predicted by the BAGEL4 server.
These include bottromycin, sactipeptides, carocin D, and colicin E9 as
shown in Table 4. The identication of RiPPs and bacteriocins, including
bottromycin, sactipeptides, carocin D, and colicin E9, provides another
layer of complexity to the potential role of M. phragmitis PSU-3885–11 in
human infections. These antimicrobial peptides may confer a competi-
tive advantage by inhibiting the growth of other bacteria in both envi-
ronmental and host-associated settings by inhibiting the growth of
competing bacteria (Nawrocki et al., 2014; Simons et al., 2020). This
result could support the survival and persistence of M. phragmitis
PSU-3885–11.
3.7. Antimicrobial resistance gene (ARG) proles among Mangrovibacter
sp.
The identication of ARGs in Mangrovibacter sp. was performed using
the Comprehensive Antibiotic Resistance Database (CARD), as illus-
trated in Fig. 3. The heatmap displays the ARG proles for M. phragmitis
PSU-3885–11 alongside data from other Mangrovibacter strains available
in the NCBI database, including Mangrovibacter sp. MFB070,
M. phragmitis MP23, M. plantisponsor DSM 19,579, and M. yixingensis
SaN21–3. The ARGs are categorized based on their roles in resistance
mechanisms including antibiotic efux, target modication, and inac-
tivation. Specically, genes involved in antibiotic efux include acrA,
acrAB-TolC, CRP, qacG, emrR, emrB, rsmA, and msbA. Genes associated
with target alteration, such as H
–
NS, arnT, PBP3, EF-Tu, and lmoG,
modify the antibiotic target sites, while only fosA8 is implicated in
Fig. 2. Distribution of MGE features in M. phragmitis PSU-3885–11. Genes are
categorized into ve functional groups, each color-coded: Integration and
excision (IE; yellow), transfer (T; light blue), replication, recombination, and
repair (RRR; dark pink), phage (P; light green), and stability, transfer, or de-
fense (STD; purple).
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Current Research in Microbial Sciences 8 (2025) 100350
5

antibiotic inactivation. For M. phragmitis PSU-3885–11, the ARG prole
indicates a range of resistance mechanisms, showing moderate to high
identity in several antibiotic efux genes such as acrA, acrAB-tolC, CRP,
and rsmA. Moreover, only the PSU-3885–11 carries msbA which can
provide this strain resistance to the nitroimidazole antibiotic class. The
antimicrobial susceptibility pattern of PSU-3885–11 indicates resistance
to beta-lactam antibiotics, such as ampicillin and cefotaxime, while
retaining susceptibility to a broad range of other antibiotics. This
resistance is likely mediated by specic ARGs identied in the genome,
including those associated with efux pumps, such as acrA, acrAB-TolC,
CRP, and rsmA. These genes are known to contribute to multidrug
resistance by actively expelling antibiotics from bacterial cells, thereby
lowering intracellular drug concentrations and diminishing their effec-
tiveness (Gaurav et al., 2023; Weston et al., 2018). Moreover, the
presence of the msbA which is associated with resistance to nitro-
imidazole antibiotics is unique in M. phragmitis PSU-3885–11 among the
studied Mangrovibacter strains (Alfaray et al., 2023). The result suggests
that this strain may possess a broader resistance prole than others.
Therefore, the antimicrobial susceptibility prole suggests the presence
of intrinsic or acquired resistance mechanisms that enable PSU-3885–11
to survive specic antibiotic treatments, especially beta-lactams, which
are commonly used to treat bacterial infections.
3.8. Virulence factor identication in Mangrovibacter sp. among available
strains
In this study, the presence and identity of key virulence factor genes
were assessed in various strains of Mangrovibacter sp. The heatmap in
Fig. 4 illustrates the conservation of selected virulence genes across four
strains. Among the genes analyzed, ompA and hcp/tssD exhibited near-
complete conservation with 100 % identity across all strains, indi-
cating a high degree of functional preservation. The gene rpoS also
showed high conservation, suggesting its importance in adapting to
hostile environments. In contrast, the genes fur and rcsB showed variable
identity across Mangrovibacter strains and exhibited lower conservation
of these genes.
The detection of genes associated with virulence, stress response, and
host adaptation, such as ompA, hcp/tssD, and rpoS, suggests that PSU-
3885–11 possesses a genetic repertoire capable of survival in hostile
environments, including the human host. The ompA, encoded for outer
membrane protein A, plays a critical role in adhesion to host cells,
promoting colonization and invasion. This adhesion ability is critical for
the establishment of infections, as it allows bacteria to interact inti-
mately with host tissues, a key step in the pathogenesis process. More-
over, ompA has been extensively reported as a major virulence factor
across a wide range of human opportunistic pathogens, including Aci-
netobacter baumannii, Acinetobacter nosocomialis, Pseudomonas
Table 3
Restriction-Modication sites identied in M. phragmitis PSU-3885–11.
Gene %Identity Query length Contig Position Type Function Accession No.
EcoEI 88.29 2442 / 2442 3885–11_00008 101,254……103,695 Type I restriction enzyme NEBM62
M.Ecl93I 88.59 1507 / 1512 3885–11_00013 122,784…124,290 Type I methyltransferase CP027604
M.Ecl884AI 90.97 1473 / 1473 3885–11_00008 99,709……101,181 Type I methyltransferase CP022532
Table 4
Bacteriocins identied M. phragmitis PSU-3885–11.
Contig Start End Class E-value
Contig 1 686,638 697,258 Colicin E9 9.56e-26
Contig 5 135,050 155,050 Bottromycin 1.96e-18
Contig 14 54,623 76,219 Sactipeptides 1.05e-27
Contig 14 89,681 110,053 Carocin D 8.44e-178
Fig. 3. Antimicrobial resistance gene (AMR) prole of Mangrovibacter sp. across available strains from the NCBI database. The percentage identications of the gene
are represented by colors.
Fig. 4. Heatmap of virulence factor genes identied in Mangrovibacter sp. The heatmap displays the percentage of virulence-related genes across one strain in this
study and the other four isolates obtained from the NCBI database. The color gradient represents the percentage identity, ranging from 95 % to 100 %, with darker
red indicating higher identity.
N. Chaichana et al.
Current Research in Microbial Sciences 8 (2025) 100350
6

aeruginosa, Escherichia coli, Cronobacter sakazakii, and Salmonella
Typhimurium (Gao et al., 2021; Kim et al., 2016; Paulsson et al., 2021;
Roy Chowdhury et al., 2022). In these pathogens, ompA contributes not
only to adhesion but also to biolm formation, immune evasion, and
intracellular survival, which are essential for persistence and infection
progression. Similarly, hcp/tssD, a key component of the Type VI
secretion system (T6SS), enables the transport of effector proteins be-
tween neighboring cells. The T6SS functions by injecting toxic effector
proteins directly into competing bacterial cells, providing a competitive
edge and possibly facilitating interactions with host cells to enhance
infection (Hernandez et al., 2020; Ma et al., 2017). Previous studies have
extensively highlighted the dual role of the T6SS in both microbial
competition and host-pathogen interactions (Gallegos-Monterrosa and
Coulthurst, 2021; Yin et al., 2024). For example, T6SS-mediated de-
livery of antibacterial toxins, such as cell wall-degrading enzymes, has
been shown to provide a signicant advantage in polymicrobial envi-
ronments by eliminating competing bacteria (Alcoforado Diniz et al.,
2015). In P. aeruginosa, T6SS has been shown to deliver effector proteins
into prokaryotic and eukaryotic cells to enhance the survival of the
donor cell (Wood et al., 2019). The near-complete conservation of ompA
and hcp/tssD across various Mangrovibacter strains highlights their
essential roles in bacterial survival and pathogenicity by maintaining
cell envelope integrity, facilitating bacterial interactions, and helping
bacteria evade the immune system by interacting with host complement
proteins (Confer and Ayalew, 2013; Hersch et al., 2020). Furthermore,
rpoS acts as a global regulator of stress response, activating genes that
enable bacterial survival under harsh conditions such as oxidative stress,
nutrient deprivation, acid stress, osmotic stress, exposure to antimicro-
bial agents, and immune attacks. By enabling bacterial adaptation to
such challenges, rpoS plays a pivotal role in ensuring long-term persis-
tence, particularly in chronic infections (da Cruz Nizer et al., 2021; Zhu
et al., 2024). In E. coli, rpoS has been shown to enhance resistance to
oxidative bursts by activating the expression of katE, a catalase gene
crucial for neutralizing hydrogen peroxide (Fasnacht and Polacek,
2021). Moreover, the role of rpoS extends beyond stress tolerance to
broader survival strategies, including biolm formation, quorum
sensing regulation, and virulence factor expression in pathogens (Zhang
et al., 2021). The high conservation of rpoS suggests its vital role in the
stress response, which could be advantageous for survival under unfa-
vorable conditions, such as in a human host with a compromised im-
mune system. Interestingly, the variable conservation of genes such as
fur and rcsB, which are associated with iron regulation and biolm
formation, respectively, across different Mangrovibacter strains, suggests
strain-specic adaptations or reduced reliance on these pathways for
virulence (Yuan et al., 2020). The fur is encoded for a regulator of iron
homeostasis which might have evolved alternative strategies to acquire
iron or may inhabit niches where iron is more readily available,
reducing the need for strict iron regulation (Latorre et al., 2018). Pre-
vious studies have shown that fur regulates the abundance of key viru-
lence factors in Staphylococcus aureus, coordinating their expression
during pathogenesis. In S. aureus pneumonia, fur adapts gene expression
to iron-limited conditions, facilitating immune evasion and infection
establishment (Torres Victor et al., 2010). Likewise, rcsB encodes the
response regulator of the rcs (regulator of capsule synthesis)
two-component regulatory system, a critical pathway involved in bio-
lm formation and other surface-associated phenotypes in bacteria.
Biolm is a key factor in chronic infections and environmental persis-
tence in bacteria (Mikkelsen et al., 2013). In chronic infections, biolm
formation regulated by rcsB contributes signicantly to bacterial sur-
vival and other critical virulence factors in pathogens (Fei et al., 2021; Li
et al., 2023; Meng et al., 2021). Overall, the identied virulence factors
may enable M. phragmitis PSU-3885–11 to adapt, survive, and effectively
interact with its host. For example, genes associated with adhesion,
biolm formation, or secretion systems could enable the bacteria to
colonize host tissues, evade immune responses, or acquire essential
nutrients. While Mangrovibacter species are primarily environmental
bacteria, the presence of such virulence-related genes raises the possi-
bility of opportunistic infections, particularly in immunocompromised
individuals or under specic environmental triggers. Therefore, recog-
nizing the functional roles of these factors in human infections is
essential for evaluating the potential risks these bacteria may pose and
their ability to adapt to host environments, emphasizing the importance
of monitoring their virulence gene proles.
By comprehensively characterizing the genome of M. phragmitis PSU-
3885–11, isolated from a patient in Thailand, we gained valuable in-
sights into its genetic features, potential pathogenicity, and antibiotic
resistance prole. Moreover, the transition from an environmental niche
to a clinical setting not only underscores its survival mechanisms but
also provides a deeper understanding of how environmental bacteria
may adapt to colonize human hosts. This study emphasizes the critical
need for ongoing surveillance and genomic analysis of environmental
bacteria that may emerge as opportunistic pathogens in clinical envi-
ronments. The presence of virulence factors, ARGs, and MGEs in
M. phragmitis PSU-3885–11 points out the need for further investigation
into its clinical signicance and potential impact on public health.
4. Conclusions
This study offers signicant insights into the genomic characteristics,
antibiotic resistance, and potential pathogenicity of M. phragmitis PSU-
3885–11, a bacterium generally associated with plant roots but isolated
from a human patient with spinal tuberculosis. The whole genome
sequencing conrmed its identity and revealed antibiotic resistance
genes, mobile genetic elements, and virulence factors, which may
facilitate its persistence in human hosts as an opportunistic pathogen.
The resistance to beta-lactam antibiotics, alongside the presence of
antimicrobial peptides and bacteriocins of this strain, suggests its
adaptability in both environmental and clinical settings. These ndings
emphasize the importance of ongoing surveillance of environmental
bacteria, which may transition to human pathogens, particularly in
immunocompromised individuals.
Author contributions
Conceptualization, N.C., K.S. (Komwit Surachat) and K.S. (Kamonnut
Singkhamanan); methodology, N.C., T.Y., S.S., M.Y., N.T., and M.W.;
software, N.C., T.Y., S.S. and K.S. (Komwit Surachat); validation, K.S.
(Komwit Surachat), N.C., T.Y. and R.P.; formal analysis, N.C. and K.S.
(Komwit Surachat); investigation, N.C. and K.S. (Komwit Surachat);
resources, M.Y., S.C. and R.P.; data curation, N.C., T.Y. and K.S. (Komwit
Surachat); writing—original draft preparation, N.C., T.Y., K.S. (Komwit
Surachat) and K.S. (Kamonnut Singkhamanan); writing—review and
editing, N.C., T.Y., K.S. (Komwit Surachat) and K.S. (Kamonnut Sing-
khamanan); visualization, N.C., T.Y. and K.S. (Komwit Surachat); su-
pervision, K.S. (Komwit Surachat) and K.S. (Kamonnut Singkhamanan);
project administration, K.S. (Komwit Surachat); funding acquisition, K.
S. (Komwit Surachat). All authors have read and agreed to the published
version of the manuscript.
Institutional review board statement
This study was conducted in accordance with the Declaration of
Helsinki and approved by the Human Research Ethics Committee
(HREC) of Prince of Songkla University (protocol code: 64–284–14–1,
date of approval: 9 June 2021).
Informed consent statement
According to retrospective reviews, the ethical committee allowed
the waiver of consent forms.
N. Chaichana et al.
Current Research in Microbial Sciences 8 (2025) 100350
7

Declaration of competing interest
The authors declare that they have no known competing nancial
interests or personal relationships that could have appeared to inuence
the work reported in this paper.
Funding
This research was supported by the NSRF via the Program Manage-
ment Unit for Human Resources & Institutional Development, Research
and Innovation, grant numbers B13F660074 and B13F670076,
respectively.
Supplementary materials
Supplementary material associated with this article can be found, in
the online version, at doi:10.1016/j.crmicr.2025.100350.
Data availability
Data will be made available on request.
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