Genomic analysis of Legionella pneumophila in the drinking water system of a large building over 25 years.

Legionella pneumophila,the causative agent of Legionnaires’ disease, is often found in the plumbing systems of buildings from where it can be transmitted to humans via inhalation or aspiration of contaminated water drops. Annual routine water sampling from the potable water system of an occupational healthcare building in Basel over 25 years was performed in accordance with national guidelines. Overall, 309 water samples were collected at 38 time points over the period of 25 years.L. pneumophilawas recovered from 120 water samples (38.8%) from 26 time points. No clinical infections were recorded during this period. Initial decontamination measures were successful for approximately 12 years, after which an increase in the total number ofLegionellacolony forming units as well as ofL. pneumophila-positive sites was noticed, in 2008. Whole genome sequencing (WGS) analysis of n=123 isolates from n=113 samples showed allL. pneumophilato be sequence type (ST)-45 (Sequence based typing scheme). The isolates are closely related, with only 408 single nucleotide polymorphisms (SNPs) among all isolates after the bioinformatic removal of recombination events. Over the 25 years, a single lineage deriving from a recent common ancestor colonized the water system of this building. The phylogeny of isolate genomes can be interpreted as inferring good water circulation, possible recolonization from a common source after cleaning, with genome evolution and insertion / loss of large elements evident. Regular monitoring of waterlines in healthcare settings helps to identify concentrations ofLegionellaspp. and WGS is recommended for detailed investigation.Data SummaryAll data is submitted to the ENA under project PRJEB79004 under accession numbers ERR13662450-ERR13662572.Impact StatementThis is the most detailed, long-term study ofL. pneumophilain the water system of a single building recorded to date. TheL. pneumophilaisolates found in the building over the sampling period of 25 years were all closely related, belonging to ST45. SNP analysis suggested that the common ancestor of the cluster was from around 1938 (range 1911 to 1959), and movement of a large genomic island and plasmid transfer were observed. Despite several decontamination measures, it was impossible to completely eradicateLegionellaspp. from the water system of the historic building. No infections could be attributed to the presence ofL. pneumophilain this building. To mitigate the risk of Legionellosis from such buildings, awareness, regular water testing based on official national guidelines and recommendations, and other control measures, such as the use of sterile water for critical procedures can be recommended.

Newborn screening: adapting to advancements in whole-genome sequencing.

<i>Newborn Screening:</i> Adapting to Advancements in Whole-Genome Sequencing

To investigate the prevalence of culturable and nonculturable Legionella species in hot water systems of public buildings in Japan and assess the risk factors associated with Legionella contamination in hot water systems. Legionella species were detected by conventional culture and molecular methods in 130 water samples collected from 40 buildings. A total of 26 (20.0%) water samples from 17 (42.5%) buildings were positive by culture, qualitative PCR or both methods: Legionella pneumophila and Leg. anisa were detected in four samples by a culture method, whereas 23 samples were positive by qualitative PCR, with the presence of various Legionella species confirmed by sequencing. Of these 23 samples, bacterial counts were quantifiable in 21 by real-time PCR (from 1.7 x 10(5) to 2.6 x 10(11) cells per litre). Phylogenetic analysis of amplified partial 16S rRNA gene showed close relations to various species of Legionella, including Leg. anisa and Leg. micdadei, all of which have been associated with respiratory diseases or increased antibody titres in human sera. Assessment of risk factors showed that turbidity, free chlorine concentration, iron concentration and heterotrophic plate count (HPC) were significantly associated with Legionella contamination (P < 0.05). Contamination of hot water systems of public buildings with culturable and nonculturable Legionella species may be a potential risk factor for Legionella infection in Japan. Adequate levels of chlorine, low levels of iron and HPC are important maintenance measures in the reduction of Legionella contamination in hot water systems. More than 40% of hot water systems in the Japanese public buildings examined were contaminated by not only culturable Leg. pneumophila and Leg. anisa but also by nonculturable pathogenic species. To our knowledge, this is the first report of both culturable and nonculturable Legionella contamination in hot water systems of public buildings in Japan.

Vital Signs: Deficiencies in Environmental Control Identified in Outbreaks of Legionnaires’ Disease—North America, 2000–2014

Fundamental improvement was made for genome sequencing since the next‐generation sequencing (NGS) came out in the 2000s. The newer technologies make use of the power of massively‐parallel short‐read DNA sequencing, genome alignment and assembly methods to digitally and rapidly search the genomes on a revolutionary scale, which enable large‐scale whole genome sequencing (WGS) accessible and practical for researchers. Nowadays, whole genome sequencing is more and more prevalent in detecting the genetics of diseases, studying causative relations with cancers, making genome‐level comparative analysis, reconstruction of human population history, and giving clinical implications and instructions. In this review, we first give a typical pipeline of whole genome sequencing, including the lab template preparation, sequencing, genome assembling and quality control, variants calling and annotations. We compare the difference between whole genome and whole exome sequencing (WES), and explore a wide range of applications of whole genome sequencing for both mendelian diseases and complex diseases in medical genetics. We highlight the impact of whole genome sequencing in cancer studies, regulatory variant analysis, predictive medicine and precision medicine, as well as discuss the challenges of the whole genome sequencing.

Can Whole Genome and Whole Transcriptome Sequencing Replace Standard Procedures in CLL Diagnostics?

Whole exome sequencing (WES) has been extensively used in genomic research. As sequencing costs decline it is being replaced by whole genome sequencing (WGS) in large-scale genomic studies, but more comparative information on WES and WGS datasets would be valuable. Thus, we have extensively compared variant calls obtained from WGS and WES of matched germline DNA samples from 96 lung cancer patients. WGS provided more homogeneous coverage with higher genotyping quality, and identified more variants, than WES, regardless of exome coverage depth. It also called more reference variants, reflecting its power to call rare variants, and more heterozygous variants that met applied quality criteria, indicating that WGS is less prone to allelic drop outs. However, increasing WES coverage reduced the discrepancy between the WES and WGS results. We believe that as sequencing costs further decline WGS will become the method of choice even for research confined to the exome.

We determined the complete genomic RNA sequence of a new type of betanodavirus Korea shellfish nervous necrosis virus (KSNNV) isolated from shellfish. Compared with other isolates representing four genotypes of betanodaviruses, the identity of the whole nucleotide sequence of the virus was in the range of 76%-83% with the presence of specific genetic motifs and formed a separate new branch in the phylogenetic analysis. In pathogenic analysis by immersion method, KSNNV-KOR1 shows 100% cumulative mortality like SFRG10/2012BGGa1 (RGNNV) in newly hatched sevenband grouper and mandarin fish, which is clearly different from those found in negative control groups. There were no significant differences in increasing rates of mortality and viral intra-tissue concentration of larval fishes infected with KSNNV-KOR1 at both 20 and 25°C water temperature. Histopathological examination of each fish species in the moribund stage revealed the presence of clear vacuoles in both brain and retinal tissues similar to typical histopathology features of RGNNV. In the present study, we first report a new betanodavirus from shellfish as the aetiological agent of viral nervous necrosis disease in fish with complete genomic nucleotide sequence and pathogenic analysis.

Details of SARS-CoV-2 reinfections at a major UK tertiary centre

Process optimization of malachite green degradation by mixed biofilm positive bacteria: Application of Box–Behnken designs, RSM and whole genome sequences analysis

BackgroundLess than two percent of the human genome is protein coding, yet that small fraction harbours the majority of known disease causing mutations. Despite rapidly falling whole genome sequencing (WGS) costs, much research and increasingly the clinical use of sequence data is likely to remain focused on the protein coding exome. We set out to quantify and understand how WGS compares with the targeted capture and sequencing of the exome (exome-seq), for the specific purpose of identifying single nucleotide polymorphisms (SNPs) in exome targeted regions.ResultsWe have compared polymorphism detection sensitivity and systematic biases using a set of tissue samples that have been subject to both deep exome and whole genome sequencing. The scoring of detection sensitivity was based on sequence down sampling and reference to a set of gold-standard SNP calls for each sample. Despite evidence of incremental improvements in exome capture technology over time, whole genome sequencing has greater uniformity of sequence read coverage and reduced biases in the detection of non-reference alleles than exome-seq. Exome-seq achieves 95% SNP detection sensitivity at a mean on-target depth of 40 reads, whereas WGS only requires a mean of 14 reads. Known disease causing mutations are not biased towards easy or hard to sequence areas of the genome for either exome-seq or WGS.ConclusionsFrom an economic perspective, WGS is at parity with exome-seq for variant detection in the targeted coding regions. WGS offers benefits in uniformity of read coverage and more balanced allele ratio calls, both of which can in most cases be offset by deeper exome-seq, with the caveat that some exome-seq targets will never achieve sufficient mapped read depth for variant detection due to technical difficulties or probe failures. As WGS is intrinsically richer data that can provide insight into polymorphisms outside coding regions and reveal genomic rearrangements, it is likely to progressively replace exome-seq for many applications.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2105-15-247) contains supplementary material, which is available to authorized users.

Clinically Significant Mutations, Deletions and Translocations Involving ETV6 Identified by Whole Genome and Whole Exome Sequencing; Report From NCI/COG Target AML Initiative

The usefulness of whole genome sequencing in the management of Staphylococcus aureus infections

Recently, three cases of Legionnaires’ disease were reported in three separate nursing centers in my geographic area, the state of New York. Because this was the first time I had seen this occur during my many years of working in long-term care, I felt this would be a good time to discuss the issue. The Centers for Disease Control and Prevention (CDC) has a tool kit for Legionnaires disease, from which this update has been derived. The website and other resources are included in this article. Legionnaires’ disease, caused by the microorganisms Legionella spp., was first described after an outbreak in 1976 among attendees at an American Legion convention in Philadelphia. The disease — a type of pneumonia — is very serious, killing about 1 in 10 people who contract it. Since 2000, cases of Legionnaires’ disease have increased fourfold in the United States. In 2016, the U.S. Department of Health and Human Services reported 6,100 cases in the United States. Because Legionnaires’ disease is frequently underdiagnosed, this may not represent the true incidence. In the natural environment, Legionella occurs in fresh water but becomes a health hazard when it invades water systems in buildings. Legionnaires’ disease and Pontiac fever — another disease caused by Legionella — are transmitted via small droplets of water in the air, which can be inhaled. It is not usually transmitted from person to person. The risk factors for Legionnaires’ disease include age 50 and older, a history of smoking, chronic lung disease, immune system disorders, systemic cancer, diabetes, renal failure, or hepatic failure. A recent overnight stay outside the home — including stay in a health care facility — is also a risk factor, as is exposure to hot tubs. Of more than 60 pathogenic species of Legionella, Legionella pneumophila serogroup 1 causes most cases of legionellosis (the collective term for Legionnaires’ disease and Pontiac fever). In natural aquatic systems, Legionella microorganisms grow and multiply within protozoa, single-celled microorganisms such as amoebae. Human alveolar macrophages resemble protozoa, which makes them a suitable host for Legionella in the human lungs. After an incubation period of two to 10 days (with an average of five to six days) from the time of exposure, the symptoms that manifest include:•Fever•Myalgia•Cough•Shortness of breath•Headache•Delirium•Radiograph positive for pneumonia The preferred diagnostic tests for Legionnaires’ disease are concurrent culture of lower respiratory secretions (e.g., sputum, bronchoalveolar lavage) on selective media and the Legionella urinary antigen test. Ideally, sputum should be obtained before starting an antibiotic, but the treatment should not be delayed to accommodate this. Urinary antigen testing is effective in detecting Legionella infections for days to weeks after treatment. Serological assays are not recommended due to their low specificity. •Legionella should be considered in patients who do not respond to antibiotics or are immunocompromised. Be aware of any recent outbreaks.•Macrolides and respiratory fluoroquinolones are the preferred agents for treatment (see “CDC Resources”).•The patient may require hospitalization.•Respiratory isolation is not required because transmission is via water droplets.•Supportive treatment, such as hydration and oxygen therapy, should be provided.•Resident and family education about disease transmission and treatment is advised.•All cases should be reported to the public health department.CDC Resources•Legionella (Legionnaires’ Disease and Pontiac Fever): Diagnosis, Treatment, and Prevention; https://www.cdc.gov/legionella/clinicians/diagnostic-testing.html•Legionella (Legionnaires’ Disease and Pontiac Fever): Guidelines, Standards, and Laws; https://www.cdc.gov/legionella/resources/guidelines.html•Toolkit: Developing a Water Management Program to Reduce Legionella Growth and Spread in Buildings; https://www.cdc.gov/legionella/wmp/toolkit/ •Legionella (Legionnaires’ Disease and Pontiac Fever): Diagnosis, Treatment, and Prevention; https://www.cdc.gov/legionella/clinicians/diagnostic-testing.html•Legionella (Legionnaires’ Disease and Pontiac Fever): Guidelines, Standards, and Laws; https://www.cdc.gov/legionella/resources/guidelines.html•Toolkit: Developing a Water Management Program to Reduce Legionella Growth and Spread in Buildings; https://www.cdc.gov/legionella/wmp/toolkit/ Proper maintenance of water systems in which Legionella may grow is key to disease prevention. The CDC encourages all owners of health care facilities to develop a water management plan aimed at reducing the risk of Legionella contamination. If Legionella is found in a health care facility’s water system, it must be eliminated. A tool kit developed by the CDC can help facilities determine the sources of contamination (see “CDC Resources”). Preventive measures such as cleaning the distal outlets and removing “dead legs” in the plumbing system (Long Term Living Contin Care Prof 2014:63:36–40) are known to be expensive and, unfortunately, ineffective. An outbreak of Legionella in a skilled nursing center is a disaster for everyone involved. It is critically important to recognize the signs and symptoms of a possible Legionella infection and to have a clinical policy/protocol in place for management. From an administrative prospective, follow the CDC guidelines for determining if the water system in your building is at risk for growing and spreading Legionella and learn about the newly published standards for Legionella water management and how to monitor and respond to changes in water quality. This should be included in the facility’s policy and procedure manual.