Economics of field size and shape for autonomous crop machines.

Field size and shape constrain spatial and temporal management of agriculture with implications for farm profitability, field biodiversity and environmental performance. Large, conventional equipment struggles to farm small, irregularly shaped fields efficiently. The study hypothesized that autonomous crop machines would make it possible to farm small, non-rectangular fields profitably, thereby preserving field biodiversity and other environmental benefits. Using the experience of the Hands Free Hectare (HFH) demonstration project, this study developed algorithms to estimate field times (h/ha) and field efficiency (%) subject to field size and shape in grain-oil-seed farms of the United Kingdom using four different equipment sets. Results show that field size and shape had a substantial impact on technical and economic performance of all equipment sets, but autonomous machines were able to farm small 1 ha rectangular and non-rectangular fields profitably. Small fields with equipment of all sizes and types required more time, but for HFH equipment sets field size and shape had least impact. Solutions of HFH linear programming model show that autonomous machines decreased wheat production cost by €15/ton to €29/ton and €24/ton to €46/ton for small rectangular and non-rectangular fields respectively, but larger 112 kW and 221 kW equipment with human operators was not profitable for small fields. Sensitivity testing shows that the farms using autonomous machines adapted easily and profitably to scenarios with increasing wage rates and reduced labour availability, whilst farms with conventional equipment struggled. Technical and economic feasibility in small fields imply that autonomous machines could facilitate biodiversity and improve environmental performance. Supplementary Information: The online version contains supplementary material available at 10.1007/s11119-023-10016-w.

Airflow accelerates bovine and human articular cartilage drying and chondrocyte death.

OBJECTIVE: Exposure of articular cartilage to static air results in changes to the extracellular matrix (ECM) and stimulates chondrocyte death, which may cause joint degeneration. However during open orthopaedic surgery, cartilage is often exposed to laminar airflow, which may exacerbate these damaging effects. We compared drying in static and moving air in terms of cartilage appearance, hydration and chondrocyte viability, and tested the ability of saline-saturated gauze to limit the detrimental effects of air exposure. DESIGN: Articular cartilage from bovine metatarsophalangeal joints (N = 50) and human femoral heads (N = 6) was exposed for 90 min to (1) static air (2) airflow (up to 0.34 m/s), or (3) airflow (0.18 m/s), covered with gauze. Following air exposure, cartilage was also rehydrated (0.9% saline; 120 min) to determine the reversibility of drying effects. The influence of airflow was assessed by studying macroscopic appearance, and quantifying superficial zone (SZ) chondrocyte viability and cartilage hydration. RESULTS: Airflow caused advanced changes to cartilage appearance, accelerated chondrocyte death, and increased dehydration compared to static air. These effects were prevented if cartilage was covered by saline-saturated gauze. Cartilage rehydration reversed macroscopic changes associated with drying but the chondrocyte death was not altered. Chondrocytes at the cut edge of cartilage were more sensitive to drying compared to cells distant from the edge. CONCLUSIONS: Airflow significantly increased articular cartilage dehydration and chondrocyte death compared to static air. As laminar airflow is routinely utilised in operating theatres, it is essential that articular cartilage is kept wet via irrigation or by covering with saline-saturated gauze to prevent chondrocyte death.

The use of hyperosmotic saline for chondroprotection: implications for orthopaedic surgery and cartilage repair.

OBJECTIVE: Articular cartilage may experience iatrogenic injury during routine orthopaedic/arthroscopic procedures. This could cause chondrocyte death, leading to cartilage degeneration and posttraumatic osteoarthritis. In an in vitro cartilage injury model, chondrocyte death was reduced by increasing the osmolarity of normal saline (NS), the most commonly-used irrigation solution. Here, we studied the effect of hyperosmolar saline (HS) on chondrocyte viability and cartilage repair in an in vivo injury model. DESIGN: Cartilage injury was induced by a single scalpel cut along the patellar groove of 8 week old rats in the absence of irrigation or with either NS (300 mOsm) or HS (600 mOsm). The percentage of cell death (PCD) within the injured area was assessed using confocal microscopy. Repair from injury was evaluated by histology/immunostaining, and inflammatory response by histology, cytokine array analysis and ELISA (enzyme-linked immunosorbent assay). RESULTS: The PCD in saline-irrigated joints was increased compared to non-irrigated (NI) joints [PCD = 20.8% (95%CI; 14.5, 27.1); PCD = 9.14% (95%CI; 6.3, 11.9); P = 0.0017]. However, hyperosmotic saline reduced chondrocyte death compared to NS (PCD = 10.4% (95%CI; 8.5, 12.3) P = 0.0024). Repair score, type II collagen and aggrecan levels, and injury width, were significantly improved with hyperosmotic compared to NS. Mild synovitis and similar changes in serum cytokine profile occurred in all operated joints irrespective of experimental group. CONCLUSIONS: Hyperosmotic saline significantly reduced the chondrocyte death associated with scalpel-induced injury and enhanced cartilage repair. This irrigation solution might be useful as a simple chondroprotective strategy and may also reduce unintentional cartilage injury during articular reconstructive surgery and promote integrative cartilage repair, thereby reducing the risk of posttraumatic osteoarthritis.

Chondrocyte death after drilling and articular screw insertion in a bovine model.

OBJECTIVE: Intra-articular screws are used for internal fixation of osteochondral fragments after fracture or osteochondritis dissecans. This causes cartilage injury potentially leading to chondrocyte death. We have visualised/quantified the hole and zone of cell death (ZCD) in cartilage after drilling/insertion of various articular screws. METHOD: Using an ex vivo bovine model with transmitted light and confocal laser scanning microscopy (CLSM), the holes and ZCD following drilling/insertion of articular screws (cortical screw, headless variable pitch metallic screw, headless variable pitch bioabsorbable screw) were evaluated. In situ chondrocyte death was determined by live/dead cell viability assay. An imaging/quantification protocol was developed to compare hole diameter and ZCD from drilling/insertion of screws into cartilage. The effect of saline irrigation during drilling on the ZCD was also quantified. RESULTS: Screw insertion created holes in cartilage that were significantly (P ≤ 0.001) less than the diameters of the equipment used. With a 1.5 mm drill, a ZCD of 580.2 ± 124 µm was produced which increased to 637.0 ± 44 µm following insertion of a 2 mm cortical screw although this was not significant (P > 0.05). The ZCD from insertion of the variable pitch headless screws (diam. 3.5 mm) was lower for the metallic compared to the bioabsorbable design (800.9 ± 159 vs 1,236.4 ± 212 µm, respectively; P < 0.01). The ZCD from drilling was reduced ∼50% (P < 0.001) by saline irrigation. CONCLUSIONS: Cartilage injury during intra-articular screw fixation caused a ZCD around the hole irrespective of screw design. Saline irrigation significantly reduced the ZCD from drilling into cartilage.

Spectral technique with convergence analysis for solving one and two-dimensional mixed Volterra-Fredholm integral equation.

A numerical approach based on shifted Jacobi-Gauss collocation method for solving mixed Volterra-Fredholm integral equations is introduced. The novel technique with shifted Jacobi-Gauss nodes is applied to reduce the mixed Volterra-Fredholm integral equations to a system of algebraic equations that has an easy solved. The present algorithm is extended to solve the one and two-dimensional mixed Volterra-Fredholm integral equations. Convergence analysis for the present method is discussed and confirmed the exponential convergence of the spectral algorithm. Various numerical examples are approached to demonstrate the powerful and accuracy of the technique.

Rapid molecular detection of tuberculosis and rifampicin drug resistance: retrospective analysis of a national U.K. molecular service over the last decade.

BACKGROUND: Fast and reliable detection of Mycobacterium tuberculosis complex (MTBC) and drug resistance is crucial in establishing effective treatment and enforcing timely public health measures. METHODS: The authors analysed the performance of a national U.K. molecular diagnostic service over a decade, based on the use of a line probe assay (Innolipa, LiPA) compared with conventional liquid and solid cultures with rapid molecular identification and culture-based drug resistance testing. FINDINGS: Data were available for 7836 consecutive patient samples using LiPA and the reference microbiological technique (conventional liquid and solid cultures with rapid molecular identification and culture-based drug resistance testing). For all sputum specimens (n=3382) the sensitivity, specificity, positive predictive value, negative predictive value and accuracy for MTBC detection were 93.4%, 85.6%, 92.7%, 86.9% and 90.7%; the equivalent values for smear-positive sputum specimens (n=2606) were 94.7%, 80.9%, 93.9%, 83.3% and 91.3%. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy for detection of rifampicin resistance in all sputum samples (n=1667) were 92.1%, 99.3%, 89.4%, 99.5% and 98.9%, respectively; the equivalent values for smear-positive sputum specimens (n=1477) were 93.3%, 99.3%, 87.5%, 99.6% and 99%. Between January 2006 and December 2008, LiPA saved 25.3 and 32.2 days for TB diagnosis and rifampicin resistance of smear-positive samples, respectively. INTERPRETATION: A molecular diagnostic service, using a non-automated line probe assay approach, provides a rapid and reliable national service for diagnosing MTBC and rifampicin resistance.

Diagnostic accuracy of the Abbott ID NOW SARS-CoV-2 rapid test for the triage of acute medical admissions.

BACKGROUND: Decisions to isolate patients at risk of having coronavirus disease 2019 (COVID-19) in the emergency department (ED) must be rapid and accurate to ensure prompt treatment and maintain patient flow whilst minimising nosocomial spread. Reverse transcription polymerase chain reaction (RT-PCR) assays are too slow to achieve this, and near-patient testing is being used increasingly to facilitate triage. The ID NOW severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) assay is an isothermal nucleic acid amplification near-patient test which targets the RNA-dependent RNA-polymerase gene. AIM: To assess the diagnostic performance of ID NOW as a COVID-19 triage tool for medical admissions from the ED of a large acute hospital. METHODS: All adult acute medical admissions from the ED between 31st March and 31st July 2021 with valid ID NOW and RT-PCR results were included. The diagnostic accuracy of ID NOW [sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV)] was calculated against the laboratory reference standard. Discrepant results were explored further using cycle threshold values and clinical data. FINDINGS: Two percent (124/6050) of medical admissions were SARS-CoV-2 positive on RT-PCR. Compared with PCR, ID NOW had sensitivity and specificity of 83.1% [95% confidence interval (CI) 75.4-88.7] and 99.5% (95% CI 99.3-99.6), respectively. PPV and NPV were 76.9% (95% CI 69.0-83.2) and 99.6% (95% CI 99.5-99.8), respectively. The median time from arrival in the ED to ID NOW result was 59 min. CONCLUSION: ID NOW provides a rapid and reliable adjunct for the safe triage of patients with COVID-19, and can work effectively when integrated into an ED triage algorithm.

Audit of laboratory diagnostic methods for syphilis in England and Wales.

OBJECTIVES: The number of cases of infectious syphilis is increasing rapidly across England and Wales. Concern has been expressed about diagnostic delay and its potential impact on patient care. A standard operating procedure for the serological diagnosis of syphilis has recently been developed by the Health Protection Agency. This study aimed to audit clinical and laboratory practice in England and Wales against this standard. METHODS: All microbiology departments, genitourinary medicine (GUM) clinics and antenatal clinics in England and Wales were invited to complete a web-based questionnaire. RESULTS: The overall response rate was 76%. Practices varied between laboratories. The proportion of microbiology departments performing enzyme immunoassay (EIA), Treponema pallidum particle agglutination assay/T pallidum haemagglutination assay, rapid plasma reagin/Venereal Disease Reference Laboratory and EIA IgM were 94%, 34%, 41% and 10%, respectively. Of these, 57% only perform a single screening assay. The turnaround time for negative results was less than 1 week for 84% of microbiology departments. For positive samples, turnaround times varied from less than 1 week to 6-8 weeks, with 19% of GUM clinics reporting turnaround times of over 3 weeks. Notably, 26% of GUM clinics and 6% of antenatal clinics reported that delays in turnaround time had adversely affected patient management in the past year. CONCLUSION: This study suggests that there is significant room to improve laboratory turnaround times for the diagnosis of syphilis in England and Wales, and such improvements would be a positive step in limiting the spread of infection and of congenital syphilis.

Manipulation under anaesthesia and early physiotherapy facilitate recovery of patients with frozen shoulder syndrome.

AIM: The aim of this study was to examine the efficacy of manipulation under anaesthesia (MUA) followed by early physiotherapy in treating frozen shoulder syndrome. METHODS: In a prospective trial conducted between 26th August 2002 and 25th June 2004 in 86 patients with frozen shoulder syndrome, MUA was performed as a day procedure. Main outcomes were Disabilities of the Arm, Shoulder and Hand (DASH) score, a visual analogue score (VAS) for pain and range of movement, which were measured preoperatively and at six weeks postprocedure. RESULTS: Fifty (58.1%) patients had complete data. The average age was 54.5 years. The mean duration of symptoms until MUA was 13 months. The mean DASH score decreased from 48.07 to 15.84 (p < 0.0005). The mean VAS reduced from 6.07 to 1.88 (p < 0.0005). Flexion improved from 104.18 to 157.56 (p < 0.0005); abduction from 70.48 to 150.00 (p < 0.0005); and external rotation from 13.88 to 45.62 (p < 0.0005). CONCLUSION: MUA combined with early physiotherapy alleviates pain and facilitates recovery of function in patients with frozen shoulder syndrome.

Does a history of cardiac disease or hypertension increase mortality following primary elective total hip arthroplasty?

Preoperative co-morbidities such as known coronary artery disease have commonly deemed a patient at 'high risk' for primary elective Total Hip Arthroplasty (THA). We prospectively collected data on 1744 patients who underwent primary elective THA between 1998 and 2004; 273 had a history of cardiac disease defined as a previous hospital admission with a diagnosis of angina pectoris or myocardial infarction; 594 patients had hypertension defined as that requiring treatment with antihypertensives. We also had data on preoperative age, sex and body mass index (BMI). There was no statistically significant increase in early mortality at three months with a history of cardiac disease or hypertension and this remained so when adjusting for the other factors in a multivariate analysis. Sex or BMI also did not have a statistically significant effect on the risk of death within three months. Increasing age was the only significant risk factor for early mortality (p<0.001). Longer-term mortality at two and five years in relation to these factors was also examined. Statistical analysis revealed that coronary history now showed a highly significant association (p<0.001) with long-term mortality in patients who survived more than three months. This remained significant (p=0.002) when adjusted for the other factors. Hypertension continued to have no effect, as did BMI. Age remained a significant risk factor and ASA was also a predictor of death, as has been previously shown. The overall long-term mortality following THAwas less than expected from the normal population, even in the subgroup with a coronary history. This study will assist clinicians when advising patients who have one of these common risk factors when seeking primary elective THA.

Thaumasiovibrio occultus gen. nov. sp. nov. and Thaumasiovibrio subtropicus sp. nov. within the family Vibrionaceae, isolated from coral reef seawater off Ishigaki Island, Japan.

Two phylogenetically distinct Vibrionaceae strains C4II189T and C4V358T isolated from reef seawater off Ishigaki Island, Japan, in 2014 were studied with advanced genome-based taxonomy approaches. All aspects of phylogenetic (16S rRNA phylogeny, MLSA), phenotypic and genetic (ANI, DDH, AAI, and the number of core genes) cohesions between the two identified species were high enough to propose them as members of a new genus within the family Vibrionaceae. Consequently, an eighth genus Thaumasiovibrio gen. nov. is proposed that contains two new species Thaumasiovibrio occultus sp. nov. strain C4II189T (=DSM 101554T=JCM 31629T) (type species) and Thaumasiovibrio subtropicus sp. nov. strain C4V358T (=DSM 101555T=JCM 31630T). Thaumasiovibrio species were phylogenetically distinct from the other Vibrionaceae species based on pyrH gene sequences. The combination of catalase negative, sensitivity to vibriostatic agent O/129, and green colony formation on TCBS for the phylogenetically affiliated strains was the diagnostic features for the current tentative identification of this genus.

Vibrio aphrogenes sp. nov., in the Rumoiensis clade isolated from a seaweed.

A novel strain Vibrio aphrogenes sp. nov. strain CA-1004T isolated from the surface of seaweed collected on the coast of Mie Prefecture in 1994 [1] was characterized using polyphasic taxonomy including multilocus sequence analysis (MLSA) and a genome based comparison. Both phylogenetic analyses on the basis of 16S rRNA gene sequences and MLSA based on eight protein-coding genes (gapA, gyrB, ftsZ, mreB, pyrH, recA, rpoA, and topA) showed the strain could be placed in the Rumoiensis clade in the genus Vibrio. Sequence similarities of the 16S rRNA gene and the multilocus genes against the Rumoiensis clade members, V. rumoiensis, V. algivorus, V. casei, and V. litoralis, were low enough to propose V. aphrogenes sp. nov. strain CA-1004T as a separate species. The experimental DNA-DNA hybridization data also revealed that the strain CA-1004T was separate from four known Rumoiensis clade species. The G+C content of the V. aphrogenes strain was determined as 42.1% based on the genome sequence. Major traits of the strain were non-motile, halophilic, fermentative, alginolytic, and gas production. A total of 27 traits (motility, growth temperature range, amylase, alginase and lipase productions, and assimilation of 19 carbon compounds) distinguished the strain from the other species in the Rumoiensis clade. The name V. aphrogenes sp. nov. is proposed for this species in the Rumoiensis clade, with CA-1004T as the type strain (JCM 31643T = DSM 103759T).

Correction: Vibrio aphrogenes sp. nov., in the Rumoiensis clade isolated from a seaweed.

[This corrects the article DOI: 10.1371/journal.pone.0180053.].

Does obesity influence the clinical outcome at five years following total knee replacement for osteoarthritis?

A total of 370 consecutive primary total knee replacements performed for osteoarthritis were followed up prospectively at 6, 18, 36 and 60 months. The Knee Society score and complications (perioperative mortality, superficial and deep wound infection, deep-vein thrombosis and revision rate) were recorded. By dividing the study sample into subgroups based on the body mass index overall, the body mass index in female patients and the absolute body-weight. The outcome in obese and non-obese patients was compared. A repeated measures analysis of variance showed no difference in the Knee Society score between the subgroups. There was no statistically-significant difference in the complication rates for the subgroups studied. Obesity did not influence the clinical outcome five years after total knee replacement.

Total knee replacement in morbidly obese patients. Results of a prospective, matched study.

The results of 41 consecutive total knee replacements performed on morbidly obese patients with a body mass index > 40 kg/m(2), were compared with a matched group of 41 similar procedures carried out in non-obese patients (body mass index < 30 kg/m(2)). The groups were matched for age, gender, diagnosis, type of prosthesis, laterality and pre-operative Knee Society Score. We prospectively followed up the patients for a mean of 38.5 months (6 to 66). No patients were lost to follow-up. At less than four years after operation, the results were worse in the morbidly obese group compared with the non-obese, as demonstrated by inferior Knee Society Scores (mean knee score 85.7 and 90.5 respectively, p = 0.08; mean function score 75.6 and 83.4, p = 0.01), a higher incidence of radiolucent lines on post-operative radiographs (29% and 7%, respectively, p = 0.02), a higher rate of complications (32% and 0%, respectively, p = 0.001) and inferior survivorship using revision and pain as end-points (72.3% and 97.6%, respectively, p = 0.02). Patients with a body mass index > 40 kg/m(2) should be advised to lose weight prior to total knee replacement and to maintain weight reduction. They should also be counselled regarding the inferior results which may occur if they do not lose weight before surgery.

Drying of open animal joints in vivo subsequently causes cartilage degeneration.

OBJECTIVES: During open orthopaedic surgery, joints may be exposed to air, potentially leading to cartilage drying and chondrocyte death, however, the long-term effects of joint drying in vivo are poorly understood. We used an animal model to investigate the subsequent effects of joint drying on cartilage and chondrocytes. METHODS: The patellar groove of anaesthetised rats was exposed (sham-operated), or exposed and then subjected to laminar airflow (0.25m/s; 60 minutes) before wounds were sutured and animals recovered. Animals were monitored for up to eight weeks and then sacrificed. Cartilage and chondrocyte properties were studied by histology and confocal microscopy, respectively. RESULTS: Joint drying caused extensive chondrocyte death within the superficial regions of cartilage. Histology of dried cartilage demonstrated a loss of surface integrity at four weeks, fibrillations at eight weeks, and an increased modified Mankin score (p < 0.001). Cartilage thickness increased (p < 0.001), whereas chondrocyte density decreased at four weeks (p < 0.001), but then increased towards sham-operated levels (p < 0.01) at eight weeks. By week eight, chondrocyte pairing/clustering and cell volume increased (p < 0.05; p < 0.001, respectively). CONCLUSIONS: These in vivo results demonstrated for the first time that as a result of laminar airflow, cartilage degeneration occurred which has characteristics similar to those seen in early osteoarthritis. Maintenance of adequate cartilage hydration during open orthopaedic surgery is therefore of paramount importance.Cite this article: Dr A. Hall. Drying of open animal joints in vivo subsequently causes cartilage degeneration. Bone Joint Res 2016;5:137-144. DOI: 10.1302/2046-3758.54.2000594.

The First Temporal and Spatial Assessment of Vibrio Diversity of the Surrounding Seawater of Coral Reefs in Ishigaki, Japan.

Coral reefs perform a major role in regulating marine biodiversity and serve as hotspot for highly dynamic and diverse microbiomes as holobionts. Corals around Ishigaki, however, are at risk due to tremendous stressors including elevation of seawater temperature, eutrophication and so on. However, no information is currently available on how Vibrio diversity fluctuates spatially and temporally due to environmental determinants in Ishigaki coral reef ecosystems. The aim of this study is to elucidate spatiotemporal Vibrio diversity dynamic at both community and population levels and to assess the environmental drivers correlated to Vibrio abundance and diversity. The Vibrio community identified based on pyrH gene phylogeny of 685 isolates from seawater directly connecting to Ishigaki coral holobionts consisted of 22 known and 12 potential novel Vibrionaceae species. The most prominent species were V. hyugaensis, V. owensii and V. harveyi followed by V. maritimus/V. variabillis, V. campbellii, V. coralliilyticus, and Photobacterium rosenbergii. The Vibrio community fluctuations, assessed by PCoA with UniFrac distance and clustering with Euclidiean distance were varied less not only by year but also by site. Interestingly, significant positive correlation was observed between rising seawater temperature and the abundance of V. campbellii (r = 0.62; P < 0.05) whereas the opposite was observed for V. owensii (r = -0.58; P < 0.05) and the C6 group of V. hyugaensis (r = -0.62; P < 0.05). AdaptML-based microhabitat differentiation revealed that V. harveyi, V. campbellii, P. rosenbergii, and V. coralliilyticus populations were less-ecologically distinctive whereas V. astriarenae and V. ishigakensis were ecologically diverse. This knowledge could be important clue for the future actions of coral conservation.

Vibrio ishigakensis sp. nov., in Halioticoli clade isolated from seawater in Okinawa coral reef area, Japan.

Five novel strains showing non-motile, alginolytic, halophilic and fermentative features were isolated from seawater samples off Okinawa in coral reef areas. These strains were characterized by an advanced polyphasic taxonomy including genome based taxonomy using multilocus sequence analysis (MLSA) and in silico DNA-DNA similarity (in silico DDH). Phylogenetic analyses on the basis of 16S rRNA gene sequences revealed that the isolates could be assigned to the genus Vibrio, however they were not allocated into any distinct cluster with known Vibrionaceae species. MLSA based on eight protein-coding genes (gapA, gyrB, ftsZ, mreB, pyrH, recA, rpoA, and topA) showed the vibrios formed an outskirt branch of Halioticoli clade. The experimental DNA-DNA hybridization data revealed that the five strains were in the range of being defined as conspecific but separate from nine Halioticoli clade species. The G+C contents of the Vibrio ishigakensis strains were 47.3-49.1mol%. Both Amino Acid Identity and Average Nucleotide Identity of the strain C1(T) against Vibrio ezurae HDS1-1(T), Vibrio gallicus HT2-1(T), Vibrio halioticoli IAM 14596(T), Vibrio neonatus HDD3-1(T) and Vibrio superstes G3-29(T) showed less than 95% similarity. The genome-based taxonomic approach by means of in silico DDH values also supports the V. ishigakensis strains being distinct from the other known Halioticoli clade species. Sixteen traits (growth temperature range, DNase and lipase production, indole production, and assimilation of 10 carbon compounds) distinguished these strains from Halioticoli clade species. The names V. ishigakensis sp. nov. is proposed for the species of Halioticoli clade, with C1(T) as the type strain (JCM 19231(T)=LMG 28703(T)).

Advanced Microbial Taxonomy Combined with Genome-Based-Approaches Reveals that Vibrio astriarenae sp. nov., an Agarolytic Marine Bacterium, Forms a New Clade in Vibrionaceae.

Advances in genomic microbial taxonomy have opened the way to create a more universal and transparent concept of species but is still in a transitional stage towards becoming a defining robust criteria for describing new microbial species with minimum features obtained using both genome and classical polyphasic taxonomies. Here we performed advanced microbial taxonomies combined with both genome-based and classical approaches for new agarolytic vibrio isolates to describe not only a novel Vibrio species but also a member of a new Vibrio clade. Two novel vibrio strains (Vibrio astriarenae sp. nov. C7T and C20) showing agarolytic, halophilic and fermentative metabolic activity were isolated from a seawater sample collected in a coral reef in Okinawa. Intraspecific similarities of the isolates were identical in both sequences on the 16S rRNA and pyrH genes, but the closest relatives on the molecular phylogenetic trees on the basis of 16S rRNA and pyrH gene sequences were V. hangzhouensis JCM 15146T (97.8% similarity) and V. agarivorans CECT 5085T (97.3% similarity), respectively. Further multilocus sequence analysis (MLSA) on the basis of 8 protein coding genes (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA, and topA) obtained by the genome sequences clearly showed the V. astriarenae strain C7T and C20 formed a distinct new clade protruded next to V. agarivorans CECT 5085T. The singleton V. agarivorans has never been included in previous MLSA of Vibrionaceae due to the lack of some gene sequences. Now the gene sequences are completed and analysis of 100 taxa in total provided a clear picture describing the association of V. agarivorans into pre-existing concatenated network tree and concluded its relationship to our vibrio strains. Experimental DNA-DNA hybridization (DDH) data showed that the strains C7T and C20 were conspecific but were separated from all of the other Vibrio species related on the basis of both 16S rRNA and pyrH gene phylogenies (e.g., V. agarivorans CECT 5085T, V. hangzhouensis JCM 15146T V. maritimus LMG 25439T, and V. variabilis LMG 25438T). In silico DDH data also supported the genomic relationship. The strains C7T also had less than 95% average amino acid identity (AAI) and average nucleotide identity (ANI) towards V. maritimus C210, V. variabilis C206, and V. mediterranei AK1T, V. brasiliensis LMG 20546T, V. orientalis ATCC 33934T, and V. sinaloensis DSM 21326. The name Vibrio astriarenae sp. nov. is proposed with C7 as the type strains. Both V. agarivorans CECT 5058T and V. astriarenae C7T are members of the newest clade of Vibrionaceae named Agarivorans.