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Draft Genome Sequence of Multidrug-Resistant Vibrio parahaemolyticus Strain PH698, Infecting Penaeid Shrimp in the Philippines

DOI:10.1128/MRA.01040-19
Authors:
Cynthia Saloma at University of the Philippines
Cynthia Saloma
Sarah Mae U. Penir
Sarah Mae U. Penir
Sarah Mae U. Penir
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Joseph Matthew R. Azanza
Joseph Matthew R. Azanza
Joseph Matthew R. Azanza
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Leobert D. dela Pena
Leobert D. dela Pena
Leobert D. dela Pena
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Abstract

The emergence of multidrug-resistant bacterial strains in diverse settings has been reported globally. In the Philippine shrimp aquaculture industry, antibiotics are used for the treatment of bacterial diseases during the production cycle. We report the draft genome of Vibrio parahaemolyticus PH698, a multidrug-resistant strain isolated from a Philippine shrimp farm.
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Draft Genome Sequence of Multidrug-Resistant Vibrio
parahaemolyticus Strain PH698, Infecting Penaeid Shrimp in
the Philippines
Cynthia P. Saloma,
a,b
Sarah Mae U. Penir,
a,b
*Joseph Matthew R. Azanza,
a
Leobert D. dela Pena,
c
Roselyn C. Usero,
d
Nikko Alvin R. Cabillon,
c
*Angela Denise P. Bilbao,
c
*Edgar C. Amar
c
a
National Institute of Molecular Biology and Biotechnology, University of the Philippines, Diliman, Quezon City, Philippines
b
Philippine Genome Center, University of the Philippines, Diliman, Quezon City, Philippines
c
Southeast Asian Fisheries Development Center, Aquaculture Department, Tigbauan, Iloilo, Philippines
d
Negros Prawn Producers Cooperative (NPPC), Bacolod City, Negros Occidental, Philippines
ABSTRACT The emergence of multidrug-resistant bacterial strains in diverse set-
tings has been reported globally. In the Philippine shrimp aquaculture industry, anti-
biotics are used for the treatment of bacterial diseases during the production cycle.
We report the draft genome of Vibrio parahaemolyticus PH698, a multidrug-resistant
strain isolated from a Philippine shrimp farm.
Antibiotics are commonly used by aquaculture farmers to prevent and treat bacte-
rial infections in shrimp (1). The range of commonly used antibiotics varies widely
between shrimp-producing countries due to different management and regulation
practices. However, on account of extensive antibiotic use, there is an observable trend
of the emergence of antibiotic-resistant strains in shrimp farms (2, 3). In the Philippines,
there is no existing report of a multidrug-resistant Vibrio parahaemolyticus strain isolated
from shrimp. Here, we describe the draft genome sequence of V. parahaemolyticus
PH698, a multidrug-resistant strain isolated from the hepatopancreas of Penaeus van-
namei shrimp in a pond with an outbreak of unknown etiology.
Shrimp samples exhibiting clinical signs of disease were preferentially collected for
dissection and homogenization of their hepatopancreas. Serial dilutions of the hepa-
topancreas homogenate in sterile seawater were plated in nutrient agar (Pronadisa)
with 2% NaCl. The colony of Vibrio parahaemolyticus strain PH698 was isolated to
generate pure bacterial cultures. The isolated strain was grown overnight in nutrient
broth (Pronadisa) with 1.5% NaCl at 30°C. Automated genomic DNA extraction was
performed with the KingFisher cell and tissue DNA kit (Thermo Fisher Scientific). A
paired-end library was prepared from the genomic DNA using the Nextera XT library
preparation kit (Illumina) and was sequenced on the Illumina MiSeq platform (2 ⫻
300 bp, MiSeq reagent kit v. 3) at the DNA Sequencing Core Facility, Philippine Genome
Center, at a coverage of ⬎80⫻.
The quality of the reads was checked and verified using FastQC (4). Adapter
sequences were removed from 2,662,434 paired-end reads using Cutadapt v. 1.18 (5).
The trimmed paired-end reads were assembled into 62 contigs (N
50
, 193,506 bp) longer
than 1,000 bp using SPAdes v. 3.13.0 (6) with k-mer sizes of 21, 33, 55, 77, 99, and 127
with mismatch correction. The largest contig has a length of 468,564 bp, and the total
assembly size is 5,342,783 bp. The G⫹C content of the draft genome sequence of the
strain is 45.5%. The assignment of strain PH698 as V. parahaemolyticus was established
when the average nucleotide identity (ANI) values between the DNA sequence of the
isolate and other publicly available strains of the species were validated to be ⬎95%
using the aniM option of calculate_ani.py (7). Annotation of the draft genome with the
Citation Saloma CP, Penir SMU, Azanza JMR,
dela Pena LD, Usero RC, Cabillon NAR, Bilbao
ADP, Amar EC. 2019. Draft genome sequence
of multidrug-resistant Vibrio parahaemolyticus
strain PH698, infecting penaeid shrimp in the
Philippines. Microbiol Resour Announc
8:e01040-19. https://doi.org/10.1128/MRA
.01040-19.
Editor Frank J. Stewart, Georgia Institute of
Technology
Copyright © 2019 Saloma et al. This is an
open-access article distributed under the terms
of the Creative Commons Attribution 4.0
International license.
Address correspondence to Cynthia P. Saloma,
cpsaloma@up.edu.ph, or Edgar C. Amar,
eamar@seafdec.org.ph.
*Present address: Sarah Mae U. Penir, Max
Planck Institute for Biophysical Chemistry,
Göttingen, Germany; Nikko Alvin R. Cabillon,
The Hebrew University of Jerusalem, Jerusalem,
Israel; Angela Denise P. Bilbao, Hain Celestial
Group, New York, New York, USA.
Received 6 September 2019
Accepted 21 October 2019
Published 21 November 2019
GENOME SEQUENCES
crossm
Volume 8 Issue 47 e01040-19 mra.asm.org 1
NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v. 4.9 (8) led to the identification
of 5,084 coding sequences (CDSs), 104 tRNAs, 13 rRNAs, and 4 noncoding RNAs
(ncRNAs).
Detection of antibiotic resistance genes from the sequences in the Antibiotic Resistance
Gene-ANNOTation (ARG-ANNOT) database (9) in the raw reads of the strain was
executed using the default parameters of SRST2 v. 0.20 (10). V. parahaemolyticus strain
PH698 contains 8 antibiotic resistance genes, namely, aac3iv,ant3ia,dfra17,erea,
aph33ib,aph6id,sul2, and tetc. Altogether, the genes confer resistance for different
antibiotic classes, ranging from aminoglycosides (e.g., apramycin, dibekacin, gentami-
cin, netilmicin, sisomicin, spectinomycin, streptomycin, and tobramycin), trimethoprim,
macrolide-lincosamide-streptogramin (e.g., erythromycin), sulfonamides, and tetracy-
clines.
Data availability. The draft genome sequence and annotation of V. parahaemolyti-
cus PH698 have been deposited in GenBank under accession no. VSBR00000000 and
assembly accession no. GCF_008271865. Illumina reads have been deposited in the SRA
under accession no. SRR9870108.
ACKNOWLEDGMENTS
This research was conducted under the Philippine Shrimp Pathogenomics Program
and funded by the Department of Science and Technology-Philippine Council for
Agriculture, Aquatic, Natural Resources Research and Development (PCAARRD).
We acknowledge the help of the regional offices of the Bureau of Fisheries and
Aquatic Resources (BFAR) in the biosurveillance of shrimp disease outbreaks in the
Philippines.
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