ABSTRACT
Acinetobacter sp. PFS20 was isolated from the intestine of a farm-raised catfish. The strain exhibits resistance to cefotaxime and carries six resistance genes: OXA-1003, ADC-135, adeF, AbaQ, AmvA, and abeS. Genomic analysis predicts its pathogenic potential to humans and reveals genes encoding type I, II, and VI secretion systems.
KEYWORDS: Acinetobacter, antimicrobial resistance, catfish
ANNOUNCEMENT
Antimicrobial resistance (AMR) is a growing threat to catfish aquaculture, limiting the effectiveness of treatments for bacterial infections (1, 2). Acinetobacter species are notable for their ability to acquire and spread multidrug resistance and for their opportunistic pathogenicity in humans and animals (3). Here, we report an antimicrobial-resistant Acinetobacter sp. strain from farmed catfish in Mississippi, USA, predicted to be pathogenic to humans.
Acinetobacter sp. PFS20 was isolated from the intestine of a hybrid catfish (Ictalurus furcatus ×Ictalurus punctatus) raised without antibiotic treatment on a commercial farm. A sterile cotton swab was used to scrape the intestinal mucosa, which was then suspended in 1 mL of Luria-Bertani (LB) broth (Becton, Dickinson and Company, USA). From this suspension, 100 µL was plated onto MacConkey agar containing 4 µg/mL cefotaxime and incubated overnight at 37°C (4). Genomic DNA was extracted from an overnight culture grown at 37°C and 200 rpm shaken in 5 mL of LB broth using the DNeasy Blood & Tissue Kits (Qiagen, USA). Genomic DNA was fragmented to approximately 5 kb with a Covaris g-Tube (Covaris, USA). The library was prepared using the Native Barcoding Expansion 96 Kit V14 and Ligation Sequencing Kit V14 and sequenced on a PromethION sequencer with a R10.4.1. flow cell (Oxford Nanopore Technology, UK).
Raw sequencing data were basecalled using Dorado v.0.8.3 in the super-accurate mode, yielding 109,184 reads (N50 = 5,771 bp). Adapter sequences and small fragments (< 1 kb) were removed using Porechop_ABI v.0.5.0 (5) and chopper v.0.10.0 (6), respectively. Filtered reads were assembled with Flye v.2.9.6 (7) and polished using Racon v.1.5.0 (8). Assembly completeness and contamination were assessed with CheckM v.1.2.3 (9). Taxonomic classification was performed using GTDB-TK v2.3.0 (10) and confirmed through average nucleotide identity (ANI) using FastANI v.1.3.4 (11). Genome annotation was conducted using NCBI’s Prokaryotic Genome Annotation Pipeline (PGAP) (12). Antimicrobial resistance genes (ARGs) were identified using the Resistance Gene Identifier v.6.0.5 against the CARD 4.0.1 database (13). The pathogenic potential was predicted using PathogenFinder 1.1 (14). Default parameters were used for the software except where otherwise noted.
The key genomic characteristics of Acinetobacter sp. PFS20 are summarized in Table 1. The draft genome was assembled into two contigs, totaling approximately 4.00 Mbps, and 99.94% genome completeness (0.36% contamination). Acinetobacter sp. PFS20 showed the highest genomic similarity to an indole-degrading strain, Acinetobacter sp. JW (15) (ANI = 96.91%), followed by a diesel-degrading strain, Acinetobacter oleivorans DR1 (ANI = 95.36%) (16). Acinetobacter sp. PFS20 encodes six antimicrobial resistance genes, including OXA-1003 (OXA β-lactamase), ADC-135 (ADC β-lactamase with unclassified carbapenemase activity), and four efflux pump genes (adeF, Acinetobacter baumannii AbaQ, Acinetobacter baumannii AmvA, and abeS). Moreover, it is predicted to be a human pathogen (probability: 84.8%) and harbors genes encoding type I, II, and VI secretion systems.
TABLE 1.
Summary of sequence data and genomic features of the Acinetobacter sp. PFS20
| Feature | Value |
|---|---|
| Number of basecalled reads | 109,184 |
| N50 of basecalled reads (bp) | 5,771 |
| Genome size (bp) | 3,998,343 |
| Contig count | 2 |
| Contig N50 (bp) | 3,997,781 |
| Coverage depth (sequencing depth) | 112× |
| Completeness (%) | 99.94 |
| Contamination (%) | 0.36 |
| Guanine-cytosine content (%) | 38.82 |
| Number of genes | 3,780 |
| Number of coding sequences (CDSs) | 3,685 |
| Number of tRNAs | 73 |
| Number of rRNAs | 18 |
ACKNOWLEDGMENTS
This research was funded by the USDA‐ARS sponsored project of the Mississippi Center for Enhancing Utilization and Safety of Catfish and Other Aquatic Foods.
Contributor Information
Peixin Fan, Email: pf324@msstate.edu.
Frank J. Stewart, Montana State University, Bozeman, Montana, USA
DATA AVAILABILITY
The genome sequence of Acinetobacter sp. PFS20 and its raw sequences are available in GenBank under the accession number JBPVZE000000000 and in the Sequence Read Archive under accession number SRR34662555, respectively.
ETHICS APPROVAL
The catfish utilized in this study were obtained from a commercial aquaculture facility, where they were harvested and slaughtered as part of routine production operations.
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Associated Data
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Data Availability Statement
The genome sequence of Acinetobacter sp. PFS20 and its raw sequences are available in GenBank under the accession number JBPVZE000000000 and in the Sequence Read Archive under accession number SRR34662555, respectively.