RNA-seq is an effective method for studying the transcriptome, but it can be difficult to apply to scarce or degraded RNA from fixed clinical samples, rare cell populations or cadavers. Recent studies have proposed several methods for RNA-seq of low-quality and/or low-quantity samples, but the relative merits of these methods have not been systematically analyzed. Here we compare five such methods using metrics relevant to transcriptome annotation, transcript discovery and gene expression. Using a single human RNA sample, we constructed and sequenced ten libraries with these methods and compared them against two control libraries. We found that the RNase H method performed best for chemically fragmented, low-quality RNA, and we confirmed this through analysis of actual degraded samples. RNase H can even effectively replace oligo(dT)-based methods for standard RNA-seq. SMART and NuGEN had distinct strengths for measuring low-quantity RNA. Our analysis allows biologists to select the most suitable methods and provides a benchmark for future method development.

/pdf/comparative-analysis-of-rna-sequencing-methods-for-degraded-1n5xicmlz8.pdf

Comparative analysis of RNA sequencing methods for degraded or low-input samples

Plant–microbe interactions in the rhizosphere have important roles in biogeochemical cycling, and maintenance of plant health and productivity, yet remain poorly understood. Using RNA-based metatranscriptomics, the global active microbiomes were analysed in soil and rhizospheres of wheat, oat, pea and an oat mutant (sad1) deficient in production of anti-fungal avenacins. Rhizosphere microbiomes differed from bulk soil and between plant species. Pea (a legume) had a much stronger effect on the rhizosphere than wheat and oat (cereals), resulting in a dramatically different rhizosphere community. The relative abundance of eukaryotes in the oat and pea rhizospheres was more than fivefold higher than in the wheat rhizosphere or bulk soil. Nematodes and bacterivorous protozoa were enriched in all rhizospheres, whereas the pea rhizosphere was highly enriched for fungi. Metabolic capabilities for rhizosphere colonisation were selected, including cellulose degradation (cereals), H2 oxidation (pea) and methylotrophy (all plants). Avenacins had little effect on the prokaryotic community of oat, but the eukaryotic community was strongly altered in the sad1 mutant, suggesting that avenacins have a broader role than protecting from fungal pathogens. Profiling microbial communities with metatranscriptomics allows comparison of relative abundance, from multiple samples, across all domains of life, without polymerase chain reaction bias. This revealed profound differences in the rhizosphere microbiome, particularly at the kingdom level between plants.

/pdf/comparative-metatranscriptomics-reveals-kingdom-level-2kdujc3zo0.pdf

Comparative metatranscriptomics reveals kingdom level changes in the rhizosphere microbiome of plants

Outbreaks of Middle East respiratory syndrome (MERS) raise questions about the prevalence and evolution of the MERS coronavirus (CoV) in its animal reservoir. Our surveillance in Saudi Arabia in 2014 and 2015 showed that viruses of the MERS-CoV species and a human CoV 229E-related lineage co-circulated at high prevalence, with frequent co-infections in the upper respiratory tract of dromedary camels. viruses of the betacoronavirus 1 species, we found that dromedary camels share three CoV species with humans. Several MERS-CoV lineages were present in camels, including a recombinant lineage that has been dominant since December 2014 and that subsequently led to the human outbreaks in 2015. Camels therefore serve as an important reservoir for the maintenance and diversification of the MERS-CoVs and are the source of human infections with this virus.

https://science.sciencemag.org/content/sci/351/6268/81.full.pdf

Co-circulation of three camel coronavirus species and recombination of MERS-CoVs in Saudi Arabia

The first known cases of Middle East respiratory syndrome (MERS), associated with infection by a novel coronavirus (CoV), occurred in 2012 in Jordan but were reported retrospectively. The case first to be publicly reported was from Jeddah, in the Kingdom of Saudi Arabia (KSA). Since then, MERS-CoV sequences have been found in a bat and in many dromedary camels (DC). MERS-CoV is enzootic in DC across the Arabian Peninsula and in parts of Africa, causing mild upper respiratory tract illness in its camel reservoir and sporadic, but relatively rare human infections. Precisely how virus transmits to humans remains unknown but close and lengthy exposure appears to be a requirement. The KSA is the focal point of MERS, with the majority of human cases. In humans, MERS is mostly known as a lower respiratory tract (LRT) disease involving fever, cough, breathing difficulties and pneumonia that may progress to acute respiratory distress syndrome, multiorgan failure and death in 20 % to 40 % of those infected. However, MERS-CoV has also been detected in mild and influenza-like illnesses and in those with no signs or symptoms. Older males most obviously suffer severe disease and MERS patients often have comorbidities. Compared to severe acute respiratory syndrome (SARS), another sometimes- fatal zoonotic coronavirus disease that has since disappeared, MERS progresses more rapidly to respiratory failure and acute kidney injury (it also has an affinity for growth in kidney cells under laboratory conditions), is more frequently reported in patients with underlying disease and is more often fatal. Most human cases of MERS have been linked to lapses in infection prevention and control (IPC) in healthcare settings, with approximately 20 % of all virus detections reported among healthcare workers (HCWs) and higher exposures in those with occupations that bring them into close contact with camels. Sero-surveys have found widespread evidence of past infection in adult camels and limited past exposure among humans. Sensitive, validated reverse transcriptase real-time polymerase chain reaction (RT-rtPCR)-based diagnostics have been available almost from the start of the emergence of MERS. While the basic virology of MERS-CoV has advanced over the past three years, understanding of the interplay between camel, environment, and human remains limited.

/pdf/mers-coronavirus-diagnostics-epidemiology-and-transmission-138610lrnv.pdf

MERS coronavirus: diagnostics, epidemiology and transmission

High throughput sequencing methods and analysis for microbiome research

Next-generation sequencing has great potential for application in bacterial transcriptomics. However, unlike eukaryotes, bacteria have no clear mechanism to select mRNAs over rRNAs; therefore, rRNA removal is a critical step in sequencing-based transcriptomics. Duplex-specific nuclease (DSN) is an enzyme that, at high temperatures, degrades duplex DNA in preference to single-stranded DNA. DSN treatment has been successfully used to normalize the relative transcript abundance in mRNA-enriched cDNA libraries from eukaryotic organisms. In this study, we demonstrate the utility of this method to remove rRNA from prokaryotic total RNA. We evaluated the efficacy of DSN to remove rRNA by comparing it with the conventional subtractive hybridization (Hyb) method. Illumina deep sequencing was performed to obtain transcriptomes from Escherichia coli grown under four growth conditions. The results clearly showed that our DSN treatment was more efficient at removing rRNA than the Hyb method was, while preserving the original relative abundance of mRNA species in bacterial cells. Therefore, we propose that, for bacterial mRNA-seq experiments, DSN treatment should be preferred to Hyb-based methods.

/pdf/duplex-specific-nuclease-efficiently-removes-rrna-for-3ft4g6t9yy.pdf

Duplex-specific nuclease efficiently removes rRNA for prokaryotic RNA-seq

The full genome sequence of a Middle East respiratory syndrome coronavirus (MERS-CoV) was identified from cultured and isolated in Vero cells. The viral genome sequence has high similarity to 53 human MERS-CoVs, ranging from 99.5% to 99.8% at the nucleotide level.

/pdf/complete-genome-sequence-of-middle-east-respiratory-syndrome-9rmvq36azf.pdf

Complete Genome Sequence of Middle East Respiratory Syndrome Coronavirus KOR/KNIH/002_05_2015, Isolated in South Korea.

Global efforts have markedly decreased the disease burden of vaccine-preventable diseases. Many countries have made considerable progress toward the elimination of measles. As elimination is approached, the very low incidence achieved by high vaccination coverage has underscored the need for a sensitive and timely surveillance system. In the Republic of Korea, an active laboratory surveillance system (ALSS) was implemented to supplement the existing passive surveillance system in 2006. The ALSS connects 5 major commercial laboratories and the national measles reference laboratory, where referred samples with positive or equivocal results are retested. Annually, from 2009 to 2013, 3714 suspected cases were detected through the ALSS, an expansion of 8- to 57-fold, compared with only the passive surveillance system. The ALSS, with its sensitivity and timeliness, is a reasonable strategy to supplement the existing measles surveillance system and to help identify the elimination of measles.

How Can We Identify the Elimination of Infectious Diseases? Experience From an Active Measles Laboratory Surveillance System in the Republic of Korea.

ESG 경영의 필요성은 날이 갈수록 증가하고 있다. 항공사는 탄소 배출 및 산업 안전 문제의 중심에서 ESG 경영의 필요성을 요구받고 있다. 본 연구의 목적은 ESG 경영 활동을 토대로 한 홍보 전략과 일반적인 가격 전략의 차이를 비교하여 항공사의 서비스 수준에 따라 보다 효과적인 홍보 전략을 수립하는 데 제언하는 것으로 실험을 통해 총 네 가지 차원의 시나리오를 구성하여 연구를 진행하였다. 연구 진행의 전제는 다음과 같다. 첫째, 프리미엄 항공사인 FSC의 주 고객은 가격에 상대적으로 둔감하여 무조건 싼 가격의 항공권을 구매하기보다는 자신의 가치관 등 기타 가격 이외의 요소에 많은 관심을 두고 있을 것이고 저가 항공사인 LCC의 주 고객은 상대적으로 가격에 민감하여 구매 이전의 정보 탐색 과정에서 준거 가격 이하의 항공권을 찾는 데 많은 시간을 할애할 것이다. 둘째, 이러한 차이는 항공사가 제공하는 서비스의 수준과 항공권의 가격 차이에서 야기되며 이 차이는 인지적, 정서적, 행동적 인게이지먼트의 세 가치 차원의 차이를 야기할 것이다.BR 분석 결과, FSC는 ESG 경영 활동을 홍보 게시물의 내용으로 구성하였을 때, 인지적, 정서적, 행동적 인게이지먼트가 높은 것으로 나타났다. LCC는 할인 프로모션 등의 가격 전략을 홍보 게시물을 내용으로 구성하였을 때 인지적, 정서적, 행동적 인게이지먼트가 높은 것으로 나타났다. 본 연구의 결과에 따르면 소셜미디어를 활용한 항공사의 홍보 전략 수립은 항공사가 제공하는 서비스 수준의 차이를 고려해야 한다. 즉, 항공사의 홍보 전략 수립은 주 고객층의 특성 차이를 고려한 수준에서 수립되어야 하며 이 과정이 제대로 수행되었을때 소비자의 인게이지먼트가 높다는 것을 확인할 수 있었다. 

Su Jin Kim

Papers

Duplex-specific nuclease efficiently removes rRNA for prokaryotic RNA-seq

Complete Genome Sequence of Middle East Respiratory Syndrome Coronavirus KOR/KNIH/002_05_2015, Isolated in South Korea.

How Can We Identify the Elimination of Infectious Diseases? Experience From an Active Measles Laboratory Surveillance System in the Republic of Korea.

A Study on the Differences in Engagement Due to Promotion of ESG and Pricing Strategy in Airlines