Combining second and third generation sequencing to unlock the evolutionary path of a super bacteria-Enterococcus

Enterococcus faecalis is a Gram-positive symbiotic bacteria that inhabits the human gastrointestinal tract.

Like other species in the genus Enterococcus, Enterococcus faecalis is also present in healthy humans.
It can cause life-threatening infections, especially in the hospital environment.
The natural high level of antibiotic resistance in Enterococcus faecalis is The patient is a potential threat.

Enterococcus faecalis can cause endocarditis and sepsis, urinary tract infection (UTI), meningitis and other infections in most healthy individuals.

This bacterium has a strong adaptability to chemicals and antibiotics.
With the development of time, Enterococcus faecalis has a high probability of evolving into super bacteria.

Therefore, constructing a complete bacterial evolution map and DNA sequence can help hospitals conduct careful screening, thereby reducing the possibility of infection.

A recent article entitled "Apparent nosocomial adaptation of Enterococcus faecalis predates the modern hospital era" published in Nature Communication magazine elaborated on the evolution of the bacteria.

Detailed interpretation of the following: Articles: Apparent nosocomial adaptation of Enterococcus faecalis predates the modern hospital era Author: Anna K.
Pöntinen; Janetta Top etc.
Unit: University of Oslo, Norway; University Medical Center Utrecht, Netherlands etc.
Time: 2021.
3.
9 Journal: Nature Communication Abstract Enterococcus faecalis is a common pathogen that exists in animals and insects, which threatens people's health.

In order to construct a complete evolutionary tree of bacteria, the researchers extracted a large number of samples of the bacteria from all over the world.

Bacterial sample hosts include wild birds, mammals, healthy people and hospitalized patients.

The researchers used short-read and long-read sequencing technologies to identify multiple existing hospital-related bacterial lineages (lineages), and finally discovered that the bacteria's earliest common ancestors can be traced back to the 19th century.

Through a large number of comparative analyses, it was found that although Enterococcus faecalis has a small genome, it has strong metabolic flexibility and stable core genome.

So far, a study covering a large number of bacterial samples from 1936 to 2018 has been released.

The results of the study indicate that the resistance of Enterococcus faecalis may be far earlier than the "modern medical era".

Materials and methods WGS (whole-genome sequencing) 1) Short-read sequencing: use the Illumina HiseqX platform to perform next-generation sequencing of 2027 Enterococcus faecalis isolates; use Kraken v.
0.
10.
6 to identify potential mixed strains and eliminate species contamination ; According to the genus-spcific RefSeq database, Velvet assembler and Prokka are used for genome assembly and annotation; 2) Nanopore sequencing and hybrid assembly: In order to establish the complete chromosomes and plasmids of the main lineage, 408 isolated samples are used for ONT sequencing.

Among them, 42 full-gene samples of bacteria before 1989 were labeled.

The machine-learning classifier uses the labeled information to classify the remaining bacterial samples.

Unicycler v.
0.
4.
7 is used to display the initial assembly results of Illumina-Nextseq; bioawk v.
20110810 is used to calculate the genome size of each initially assembled isolate; Filtlong v.
0.
2.
0 uses Illumina NextSeq reads to evaluate the quality of ONT Reads.

Finally, a total of 335 completely continuous contigs were assembled.

3) Phylogenetic tree In 2027 samples of Enterococcus faecalis, the core genome was determined to have 2,906,573 base pairs and 194,831 SNPs.

Then RaxML was used to construct phylogenetic analyses based on the SNP data.

Main conclusions There are a total of 2027 Enterococcus faecalis samples, of which 967 are from hospital patients and 391 are from healthy people.

Among the human-related samples, 28 samples are from the pre-antibiotic era.

The remaining samples include 156 environmental samples, 130 farm animal samples, and 219 samples from food, pets and wild animals.

The final 136 samples were from wild birds.

The sample includes 24 countries, and the time span is from 1936 to 2018.

Researchers used bioinformatics technology to identify 173 main clusters (PP), of which the number of individuals in the ten largest clusters accounted for 52.
7% of the total number of individuals.

The three largest clusters (PP) overlap with the three main sequence types (ST): ST6 (PP2), ST16 (PP3), ST40 (PP1), among which ST6 is related to humans.

Each individual cluster contains a variety of ST.
The comparative distribution of isolated sources shows that the populations of Enterococcus faecalis are interconnected across different host types.

Fig.
1 Pangenome Neighbour Identification for Bacterial Populations (PANINI) 28 networks on E.
faecalis pangenome depict isolates of different origins distributed across the clade structure and embedded within the same, clearly defined extant clusters.
Enterococcus faecalis with 335 complete continuous assembly Based on the genome, it is estimated that the average genome size of Enterococcus faecalis is 2.
97Mbp, and the plasmid genome is 24.
3kbp.

Even if the bacteria come from different hosts, the genome sizes of hospital-related lineages and non-medical-related lineages are not very different.

Similarly, the study observes common clusters of bacterial samples from different countries.
The strains of these clusters are very stable and they have metal resistance genes.
In the early agricultural and medical practices, large amounts of lead and mercury were used to resist metals.
Genes evolved from this.

In general, the Enterococcus faecalis population not only possesses a stable genome size, but is also related to each other among different hosts.

This research also integrated a variety of biological information methods to construct a phylogenetic tree and gene heat map of Enterococcus faecalis.

These experimental data also show the evolution of antimicrobial resistance and virulence factors.

Finally, it was discovered that the adaptability of Enterococcus faecalis to hospitals may be earlier than the development of modern medicine, which allows us to pay attention to the versatility of hospital pathogens.

Fig 2.
Collection-wide antimicrobial resistance gene (ARG) patterns of E.
faecalis depict abundance of antimicrobial resistance (AMR), also in isolates representing widely different host types They are mixed and used to become a powerful combination technique.

"Sequencing technology can also be well applied in ecology, geography and even large time-span sample collection.

This allows us to better restore the evolutionary process and population structure of major pathogens.

" This hybrid sequencing method It is usually helpful to reveal evolutionary trends and their evolutionary results.

In this study, ONT was used to sequence 408 samples of Enterococcus faecalis, and finally 335 consecutive and complete genomes were assembled; from this, the average bacterial genome size was calculated to be 2.
97Mbp and the plasmid size was 24.
3kbp.

The data laid a solid foundation for the entire study.

The great ancient Roman poet Horace said: "A good start is half the battle.
"
Master of Science is willing to work with you to lay this solid foundation and make your bacterial research icing on the cake.
The following ONT bacterial genome sequencing promotions are now launched: About Hope Group, as a leader in the third-generation sequencing industry, Hope Group has a complete third-generation sequencing platform , Including Nanopore's high-throughput sequencing platform PromethION and PacBio's latest Sequel II platform, a strong bioinformatics analysis team, and a self-developed NextDenovo series of algorithms.
Currently, it has provided high-quality sequencing and analysis services for many scientific research institutes and units.

The three-generation single-cell full-length transcriptome sequencing platform combines the advantages of single-cell and third-generation long-read lengths, and looks forward to assisting more scientific researchers to carry out a series of scientific research projects such as developmental biology and tumor research.

Call 153 8703 7487 Email: sales-support@grandomics.
com or find the reference of a technology consultant near you: Pöntinen, AK, Top, J.
, Arredondo-Alonso, S.
, Tonkin-Hill, G.
, Freitas, AR , Novais, C.
,.
.
.
Corander, J.
(2021).
Apparent nosocomial adaptation of enterococcus faecalis predates the modern hospital era.
Nature Communications, 12(1).
doi:10.
1038/s41467-021-21749-5 Invade and delete from the Internet END