Lengthy Incubation Period Research Articles

Propensity of 21700 cylindrical cells to thermal runaway under slight overcharging cycling – A numerical study

Slight overcharge of lithium-ion batteries (LIBs) could occur due to inadequate design of battery management system or unexpected malfunction of charger. In comparison with other abuse conditions, the evolution of thermal runaway (TR) under slight overcharging often requires a lengthy incubation period, necessitating relatively long periods for experiments. Numerical simulations with validated models offer a viable alternative to evaluate such risks in modules/packs and associated mitigation measures. Since a current-cutting device is usually installed inside the LIB module/packs, the current will be cut off once the overcharge reaches a certain level, therefore, slight overcharging is of more potential relevance. However, previous simulations mainly focused on TR process induced by continuous overcharging, the conditions associated with slight overcharging have been overlooked. As slight overcharging is difficult to detect, the generated heat could easily accumulate as the cycle process prevails, result in unwanted cell temperature increases with increasing propensity for TR. A three-dimensional (3-D) predictive tool for the transition of cylindrical 21700 cell from slight overcharging cycle to TR has been developed by implementing the published models for heat generation from various thermal decomposition reactions and ISC into in-house version of the opensource computational fluid dynamics (CFD) code OpenFOAM. The code is validated with newly conducted experiments involving both normal and slight overcharging cycles. The validated model has then been used to conduct further parametric studies to investigate the effects of the overcharge voltage, current-rates and convective cooling on the propensity and run up time to TR as well as the maximum cell temperature during TR for cells with different cathode materials under different ambient temperatures. This work forms a sound basis for evaluating TR propagation in modules/packs under slight overcharging/discharging cycles to aid the development of mitigative measures to improve LIB process safety in practical applications.

Rapid identification of bloodstream infection pathogens and drug resistance using Raman spectroscopy enhanced by convolutional neural networks.

Bloodstream infections (BSIs) are a critical medical concern, characterized by elevated morbidity, mortality, extended hospital stays, substantial healthcare costs, and diagnostic challenges. The clinical outcomes for patients with BSI can be markedly improved through the prompt identification of the causative pathogens and their susceptibility to antibiotics and antimicrobial agents. Traditional BSI diagnosis via blood culture is often hindered by its lengthy incubation period and its limitations in detecting pathogenic bacteria and their resistance profiles. Surface-enhanced Raman scattering (SERS) has recently gained prominence as a rapid and effective technique for identifying pathogenic bacteria and assessing drug resistance. This method offers molecular fingerprinting with benefits such as rapidity, sensitivity, and non-destructiveness. The objective of this study was to integrate deep learning (DL) with SERS for the rapid identification of common pathogens and their resistance to drugs in BSIs. To assess the feasibility of combining DL with SERS for direct detection, erythrocyte lysis and differential centrifugation were employed to isolate bacteria from blood samples with positive blood cultures. A total of 12,046 and 11,968 SERS spectra were collected from the two methods using Raman spectroscopy and subsequently analyzed using DL algorithms. The findings reveal that convolutional neural networks (CNNs) exhibit considerable potential in identifying prevalent pathogens and their drug-resistant strains. The differential centrifugation technique outperformed erythrocyte lysis in bacterial isolation from blood, achieving a detection accuracy of 98.68% for pathogenic bacteria and an impressive 99.85% accuracy in identifying carbapenem-resistant Klebsiella pneumoniae. In summary, this research successfully developed an innovative approach by combining DL with SERS for the swift identification of pathogenic bacteria and their drug resistance in BSIs. This novel method holds the promise of significantly improving patient prognoses and optimizing healthcare efficiency. Its potential impact could be profound, potentially transforming the diagnostic and therapeutic landscape of BSIs.

Applications and advances in molecular diagnostics: revolutionizing non-tuberculous mycobacteria species and subspecies identification.

Non-tuberculous mycobacteria (NTM) infections pose a significant public health challenge worldwide, affecting individuals across a wide spectrum of immune statuses. Recent epidemiological studies indicate rising incidence rates in both immunocompromised and immunocompetent populations, underscoring the need for enhanced diagnostic and therapeutic approaches. NTM infections often present with symptoms similar to those of tuberculosis, yet with less specificity, increasing the risk of misdiagnosis and potentially adverse outcomes for patients. Consequently, rapid and accurate identification of the pathogen is crucial for precise diagnosis and treatment. Traditional detection methods, notably microbiological culture, are hampered by lengthy incubation periods and a limited capacity to differentiate closely related NTM subtypes, thereby delaying diagnosis and the initiation of targeted therapies. Emerging diagnostic technologies offer new possibilities for the swift detection and accurate identification of NTM infections, playing a critical role in early diagnosis and providing more accurate and comprehensive information. This review delineates the current molecular methodologies for NTM species and subspecies identification. We critically assess the limitations and challenges inherent in these technologies for diagnosing NTM and explore potential future directions for their advancement. It aims to provide valuable insights into advancing the application of molecular diagnostic techniques in NTM infection identification.

Cercospora leaf spot of olive in Uruguay

Cercospora Leaf Spot (CLS) of olive is an important fungal disease in Uruguay, causing severe early defoliation. Fungal isolates were obtained from olive leaves with typical CLS symptoms from Uruguayan orchards. The isolates were identified based on phenotypic characteristics and DNA sequence analyses. Infection processes under field conditions were characterized. Phylogenetic analyses confirmed that Pseudocercospora cladosporioides is the causal agent of CLS in Uruguay. Three colony morphologies were observed for isolates growing on potato dextrose agar. Mean conidium length ranged from 65.7 to 101.8 µm, and widths from 4.3 to 5.0 µm. Mean optimum growth temperature was 21.5°C (range 19.2 to 24.8°C). Under field conditions, initial CLS symptoms on leaves were observed 5 months after inoculation of cv. Arbequina plants, confirming the disease’s lengthy incubation period. This study shows that CLS as one of the most prevalent and destructive olive diseases in Uruguay, and emphasizes the importance of further research to develop efficient management of this disease.

Ferric chloride assisted QuEChERS method for separate detection of bifenazate and bifenazate-diazene in citrus fruits and its field validation.

Bifenazate is widely recognized as an effective acaricide for citrus production in various regions. Detecting both the parent compound of bifenazate and its metabolite, bifenazate-diazene, simultaneously can be challenging owing to their tendency to undergo chemical interconversion. Current methods developed for detecting bifenazate or bifenazate-diazene residues often involve lengthy incubation periods and may not effectively separate the two compounds. In this study, we developed a convenient and fast method based on a modified QuEChERS method assisted by oxidants to concurrently detect bifenazate and bifenazate-diazene. Based on preliminary analysis, it appears that ferric chloride has the ability to react with a reducing substance present in citrus, which may prevent the reduction of bifenazate-diazene. The method was validated and applied in a field trial. This work reports a novel strategy to establish a balanced 'neutral' condition to create a potential method for efficient determination of bifenazate acaricide residues in fruit matrices.

Isolation of Circulating Biomarkers for Liquid Biopsy using Immunoaffinity‐Based Stimuli‐Responsive Hybrid Hydrogel Beads

AbstractThis work presents chemically stable and biodegradable hydrogel beads for the isolation of circulating tumor cells (CTCs) and circulating exosomes in liquid biopsy. The liquid biopsy hydrogel beads (LBbeads) consisting of alginate and poly(vinyl alcohol) hydrogels show both chemical stability and stimuli‐degradable characteristics. Unlike single‐component hydrogels, this hybrid form is not easily degraded by buffers or cell culture media while its degradable characteristic remains; thus, it is useful in bio‐applications requiring multi‐step processes with various reagents and lengthy incubation periods. We applied our platform to clinical samples for isolating two promising circulating biomarkers for a liquid biopsy, CTCs and exosomes, by conjugating the hydrogel surface with anti‐EpCAM and anti‐CD63 antibodies, respectively, thus achieving 37.4 CTCs and comparable amount of exosome recovery per 1 milliliter of blood. The results show easy device‐free isolation and retrieval of CTCs and exosomes, with recovered circulating biomarkers successfully analyzed by western blot analysis and fluorescence microscopy. We believe that this simple and versatile platform enables us to isolate prominent circulating biomarkers for clinical use in cancer diagnosis.

Prion Diseases: A Unique Transmissible Agent or a Model for Neurodegenerative Diseases?

The accumulation and propagation in the brain of misfolded proteins is a pathological hallmark shared by many neurodegenerative diseases such as Alzheimer’s disease (Aβ and tau), Parkinson’s disease (α-synuclein), and prion disease (prion protein). Currently, there is no epidemiological evidence to suggest that neurodegenerative disorders are infectious, apart from prion diseases. However, there is an increasing body of evidence from experimental models to suggest that other pathogenic proteins such as Aβ and tau can propagate in vivo and in vitro in a prion-like mechanism, inducing the formation of misfolded protein aggregates such as amyloid plaques and neurofibrillary tangles. Such similarities have raised concerns that misfolded proteins, other than the prion protein, could potentially transmit from person-to-person as rare events after lengthy incubation periods. Such concerns have been heightened following a number of recent reports of the possible inadvertent transmission of Aβ pathology via medical and surgical procedures. This review will provide a historical perspective on the unique transmissible nature of prion diseases, examining their impact on public health and the ongoing concerns raised by this rare group of disorders. Additionally, this review will provide an insight into current evidence supporting the potential transmissibility of other pathogenic proteins associated with more common neurodegenerative disorders and the potential implications for public health.

Use of young plants for biological indexing of graft transmissible pathogens of citrus

In budwood certification programs, routinely used, technology-based laboratory methods for detecting known citrus pathogens need to be complemented with biological indexing to confirm the absence of known and unknown graft transmissible pathogens (GTPs). We report here a procedure for biological indexing using very young indicator plants, 75–90 days old. Assay results of traditional biological indexing and mini-plant indexing conducted for 51 quarantine accessions and 29 positives infected with known pathogens were compared. The evaluation indicated similar sensitivity of detection between the two methods. Traditional bioindexing requires premium greenhouse resources and a lengthy incubation period; the use of mini-plants for indexing can provide a viable alternative since the technique can be conducted with a fraction of the resources typically needed. In southern California, we were able to index for GTPs requiring cooler temperatures during summer by placing mini-bioindicator plants near the cooling pads in an evaporatively-cooled greenhouse. Indicator plants kept under favorable conditions expressed symptoms throughout the year. Limited greenhouse space is required due to the small size of the indicator plants, and they can be used for biological indexing within 3 months as compared to 9–12 months with traditional indicator plants. Young plants require much less space with 240 plants/m2 of bench space as compared to 30 plants/m2 using the conventional approach for biological indexing. The use of mini-plant biological indexing will enable citrus ‘clean stock’ and certification programs to function more efficiently with limited resources and facilities.

498. Impact of COVID-19 on UK Foundation Doctors: Education and Safety

BackgroundTransmission of a novel coronavirus (SARS-COV2) from China has caused a global pandemic, resulting in an unprecedented burden on healthcare systems worldwide.Frontline healthcare workers face considerable risk due to the potential for repeated exposure to infected individuals, many of whom may be asymptomatic given the lengthy incubation period of 5–10 days. With rising incidence, prevalence, and mortality rates, appropriate training and support is paramount in safeguarding the wellbeing of NHS staff and patients. Foundation doctors are among frontline staff being redeployed to areas of need. They are a vital resource within the NHS; and are uniquely positioned to support and staff medical teams.MethodsA 24 question digital survey with a mix of open-ended, multiple-choice, and rating-scale questions was distributed to the 18 UK Foundation Schools. Questions established trainee demographics and the quantity of COVID-19 training received; with focused questions relating to personal safety and support for vulnerable individuals.Results714 doctors completed the survey; 55% in FY2, 43% in FY1, and 2% above FY2 level. 74% of respondents reported receiving no formal training on COVID-19. Mean confidence was 84% in diagnosing COVID-19, and 56% in managing it. 75% did not receive formal training on the requisite safety precautions for suspected COVID-19 cases. 67% had not been fit tested and 75% of those who failed their fit test were not provided with alternative equipment. 30 doctors reported being pregnant and none had received additional support regarding protective measures. 47 doctors reported being immunocompromised and 42 of them had received no additional support in view of their increased risk. 93% of doctors had not received definitive guidance regarding the impact of this on their ARCP sign-off and progression of training.ConclusionAlarming proportions of Foundation trainees are not currently being provided with formal training or support for their personal safety during the COVID-19 outbreak. This is remediable through the urgent implementation of formal COVID-19 training, standardised across NHS trusts to ensure a consistent standard of care for the protection of trainees and patients.Disclosures All Authors: No reported disclosures

Behavioral strategies during incubation influence nest and female survival of Wild Turkeys.

Females must balance physiological and behavioral demands of producing offspring with associated expenditures, such as resource acquisition and predator avoidance. Nest success is an important parameter underlying avian population dynamics. Galliforms are particularly susceptible to low nest success due to exposure of ground nests to multiple predator guilds, lengthy incubation periods, and substantive reliance on crypsis for survival. Hence, it is plausible that nesting individuals prioritize productivity and survival differently, resulting in a gradient of reproductive strategies. Fine‐scale movement patterns during incubation are not well documented in ground‐nesting birds, and the influence of reproductive movements on survival is largely unknown. Using GPS data collected from female wild turkeys (n = 278) across the southeastern United States, we evaluated the influence of incubation recess behaviors on trade‐offs between nest and female survival. We quantified daily recess behaviors including recess duration, recess frequency, total distance traveled, and incubation range size for each nest attempt as well as covariates for nest concealment, nest attempt, and nest age. Of 374 nests, 91 (24%) hatched and 39 (14%) females were depredated during incubation. Average nest survival during the incubation period was 0.19, whereas average female survival was 0.78. On average, females took 1.6 daily unique recesses (SD = 1.2), spent 2.1 hr off the nest each day (SD = 1.8), and traveled 357.6 m during recesses (SD = 396.6). Average nest concealment was 92.5 cm (SD = 47). We found that females who took longer recess bouts had higher individual survival, but had increased nest loss. Females who recessed more frequently had lower individual survival. Our findings suggest behavioral decisions made during incubation represent life‐history trade‐offs between predation risk and reproductive success on an unpredictable landscape.

Early detection of viable Francisella tularensis in environmental matrices by culture-based PCR

BackgroundFrancisella tularensis is a fastidious, Gram-negative coccobacillus and is the causative agent of tularemia. To assess viability yet overcome lengthy incubation periods, a culture-based PCR method was used to detect early growth of the lowest possible number of F. tularensis cells. This method utilized a previously developed enhanced F. tularensis growth medium and is based on the change in PCR cycle threshold at the start and end of each incubation.ResultsTo test method robustness, a virulent Type A1 (Schu4) and B (IN99) strain and the avirulent Live Vaccine Strain (LVS) were incubated with inactivated target cells, humic acid, drinking and well water, and test dust at targeted starting concentrations of 1, 10, and 100 CFU mL− 1 (low, mid, and high, respectively). After 48 h, LVS growth was detected at all targeted concentrations in the presence of 106 inactivated LVS cells; while Schu4 and IN99 growth was detected in the presence of 104 Schu4 or IN99 inactivated cells at the mid and high targets. Early detection of F. tularensis growth was strain and concentration dependent in the presence of fast-growing well water and test dust organisms. In contrast, growth was detected at each targeted concentration by 24 h in humic acid and drinking water for all strains.ConclusionsResults indicated that the culture-based PCR assay is quick, sensitive, and specific while still utilizing growth as a measure of pathogen viability. This method can circumvent lengthy incubations required for Francisella identification, especially when swift answers are needed during epidemiological investigations, remediation efforts, and decontamination verification.

On the Possibility of Precision Control of the Linear Thermal Expansion Coefficient of Transparent Lithium-Aluminum- Silicate Sitals Near Zero Values

It is shown that the linear thermal expansion coefficient (CLTE) for sitalls based on the lithium aluminosilicate system can be precisely controlled near values close to zero in the temperature range from –80 to +500°C. It was found that heat-treatment for several hours at temperatures corresponding to the stage of nucleation of a crystalline phase makes it possible, after a lengthy incubation period, to change the sign of the CLTE of sitall from positive to negative, while preserving its sign constancy in the entire temperature interval near zero values of the CLTE.

Electronic bills of lading, blockchains and smart contracts

Abstract After a lengthy incubation period, electronic bills of lading (eBLs) are at last taking their first tentative steps in the shipping world. Systems currently in operation, however, are closed; only traders, carriers and banks who are members can participate. These closed schemes are not, therefore, true replacements for traditional paper bills of lading (pBLs), which can be used by anyone. Open eBL implementations are very difficult, which is no doubt why practical schemes so far have been closed. While blockchains and smart contracts make no difference to what is possible, they do to what is practicable. In particular, they may render feasible eBL implementations more open than those that have existed until now. They can obviate the need for distrusted central registries, and resolve the digital islands problem which can plague closed schemes. This article considers how an open eBL implementation might work. The schemes suggested here are rendered practicable by the new technology, should the industry choose to go down the eBL route. It may not so choose, at any rate in the short term, but if it does take this route, the article examines the legal obstacles that would need to be overcome. It assumes no change in the existing law, but concludes with a discussion of what lawmakers should consider, were they minded to assist.

Lattice strain evolution and load partitioning during creep of a Ni-based superalloy single crystal with rafted γ′ microstructure

In-situ neutron diffraction measurements were performed on monocrystalline samples of the Ni-based superalloy CMSX-4 during N-type γ′ raft formation under the tensile creep conditions of 1150 °C/100 MPa, and subsequently on a rafted sample under the low temperature/high stress creep conditions of 715 °C/825 MPa. During 1150 °C/100 MPa creep, the γ′ volume fraction decreased from ∼70% to ∼50%, the lattice parameter misfit was partly relieved, and the load was transferred from the creeping γ matrix to the γ′ precipitates. On cooling back to room temperature, a fine distribution of γ′ precipitates formed in the γ channels, and these precipitates were present in the 715 °C/825 MPa creep regime. Under low temperature/high stress creep, the alloy with rafted γ′ microstructure exhibited superior creep strength to the cuboidal γ′ microstructure produced following a standard heat-treatment. A lengthy creep incubation period was observed, believed to be associated with {111}〈110〉 dislocations hindering propagation of {111}〈112〉 dislocations. Following the creep incubation period, extensive macroscopic creep strain accumulated during primary creep as the γ phase yielded. Finally, the diffraction data suggest a loss of precipitate/matrix coherency in the (0k0) interfaces as creep strain accumulated.

Development and Validation of a Novel Real-time Assay for the Detection and Quantification of Vibrio cholerae.

Vibrio cholerae O1 and O139 has been known for its ability to cause epidemics. These strains produce cholera toxin which is the main cause of secretory diarrhea. V. cholerae non-O1 and non-O139 strains are also capable of causing gastroenteritis as well as septicemia and peritonitis. It has been proven that virulence factors such as T6SS, hapA, rtxA, and hlyA are present in almost all V. cholerae strains. It is imperative that viable but non-culturable cells of V. cholerae are also detected since they are also known to cause diarrhea. Thus, the aim of this study was to develop an assay that detects all V. cholerae regardless of their serotype, culturable state, and virulence genes present, by targeting the species specific conserved ompW sequence. The developed assay meets these goals with 100% specificity and is capable of detecting as low as 5.46 copy number of V. cholerae. Detection is rapid since neither lengthy incubation period nor electrophoresis is required. The assay had excellent repeatability (CV%: 0.24–1.32) and remarkable reproducibility (CV%: 1.08–3.7). Amplification efficiencies in the 89–100% range were observed. The assay is more economical than Taqman-based multiplex real-time PCR assays. Compared to other real-time assays, the ompW assay is specific and sensitive, has better repeatability and reproducibility, and is more economical.

Measurement of Microbial DNA Polymerase Activity Enables Detection and Growth Monitoring of Microbes from Clinical Blood Cultures

Surveillance of bloodstream infections (BSI) is a high priority within the hospital setting. Broth-based blood cultures are the current gold standard for detecting BSI, however they can require lengthy incubation periods prior to detection of positive samples. We set out to demonstrate the feasibility of using enzymatic template generation and amplification (ETGA)-mediated measurement of DNA polymerase activity to detect microbes from clinical blood cultures. In addition to routine-collected hospital blood cultures, one parallel aerobic blood culture was collected and immediately refrigerated until being transported for ETGA analysis. After refrigeration holding and transport, parallel-collected cultures were placed into a BACTEC incubator and ETGA time-course analysis was performed. Of the 308 clinical blood cultures received, 22 were BACTEC positive, and thus were initially selected for ETGA time course analysis. The ETGA assay detected microbial growth in all 22 parallel-positive blood cultures in less time than a BACTEC incubator and also yielded genomic DNA for qPCR-based organism identification. In summary, feasibility of detecting microbes from clinical blood culture samples using the ETGA blood culture assay was demonstrated. Additional studies are being considered towards development of clinically beneficial versions of this methodology.

Rapid and Simultaneous Detection of Mycobacterium tuberculosis Complex and Beijing/W Genotype in Sputum by an Optimized DNA Extraction Protocol and a Novel Multiplex Real-Time PCR

Rapid diagnosis and genotyping of Mycobacterium tuberculosis by molecular methods are often limited by the amount and purity of DNA extracted from body fluids. In this study, we evaluated 12 DNA extraction methods and developed a highly sensitive protocol for mycobacterial DNA extraction directly from sputa using surface-coated magnetic particles. We have also developed a novel multiplex real-time PCR for simultaneous identification of M. tuberculosis complex and the Beijing/W genotype (a hypervirulent sublineage of M. tuberculosis) by using multiple fluorogenic probes targeting both the M. tuberculosis IS6110 and the Rv0927c-pstS3 intergenic region. With reference strains and clinical isolates, our real-time PCR accurately identified 20 non-Beijing/W and 20 Beijing/W M. tuberculosis strains from 17 different species of nontuberculosis Mycobacterium (NTM). Further assessment of our DNA extraction protocol and real-time PCR with 335 nonduplicate sputum specimens correctly identified all 74 M. tuberculosis culture-positive specimens. In addition, 15 culture-negative specimens from patients with confirmed tuberculosis were also identified. No cross-reactivity was detected with NTM specimens (n = 31). The detection limit of the assay is 10 M. tuberculosis bacilli, as determined by endpoint dilution analysis. In conclusion, an optimized DNA extraction protocol coupled with a novel multiprobe multiplex real-time PCR for the direct detection of M. tuberculosis, including Beijing/W M. tuberculosis, was found to confer high sensitivity and specificity. The combined procedure has the potential to compensate for the drawbacks of conventional mycobacterial culture in routine clinical laboratory setting, such as the lengthy incubation period and the limitation to viable organisms.

Encefalitis rábica humana por mordedura de murciélago en un área urbana de Colombia

Human rabies encephalitis by a vampire bat bite in an urban area of Colombia A case of rabies encephalitis is presented in a teenaged male, which developed four months after a bat bite in the urban area of Floridablanca, Santander Province, Colombia. The complex clinical manifestations prevented the confirmation of an antemortem diagnosis, principally because of the lengthy incubation period and the absence of other similar urban cases. Despite application of several therapies, including the Milwaukee protocol, the patient died 19 days after hospital admission. The autopsy revealed a necrotic, acute, pan-encephalitis of rabies virus etiology. The test of direct immunofluorescence in brain tissue was positive, as well as the biologic test in mice. Serological tests indicated it to be an antigenic variant type 3, whose main reservoir is the hematophagous vampire bat, Desmodus rotundus. This is probably the first case of bat-induced rabies reported in an urban community of Colombia and one of the few in Latin America. Although rabies encephalitis by a bat bite is rare, the case serves as a notice to health authorities and to the medical community to be alert to this risk.

Development of batch-culture enrichment coupled to molecular detection for screening of natural and man-made environments in search of anammox bacteria for N-removal bioreactors systems

This investigation probed for anammox bacterial populations that would be suitable to start-up a biological reactor for N-removal. Samples of sludge and sediments from different environments were screened and used as inoculum for enrichment of anammox bacteria in batch cultures. Enrichments were monitored in order to detect anammox bacteria or their potential activity. Candidatus “Brocadia anammoxidans” was successfully enriched, detected, and identified in five of the twelve batch cultures. Furthermore, this organism was retrieved for the first time from a brackish environment. Wide-range primers used in several Polymerase Chain Reaction (PCR) attempts were unable to successfully amplify the 16S rRNA sequence from anammox populations, but were used to search for hypothetical ecological partners of anammox. A nested PCR approach with specific primers was also employed since conventional PCR was unable to amplify anammox DNA from the inocula of the successful enrichments. However, it was impossible to obtain optimal results with the different strategies utilized to improve PCR performance (higher annealing temperature or more specific primers), and only the primer set Amx368F-Amx820R resulted in an acceptable balance between both specificity and sensitivity. Although the enrichment process is relatively slow and requires lengthy incubation periods, it proved to be useful as the molecular analyses were not sensitive enough to detect anammox in the original samples or even after short enrichment periods. Therefore, a batch enrichment procedure coupled with molecular techniques was an appropriate approach to achieve successful inocula for starting-up anammox biological reactor.

Non-spore forming eubacteria isolated at an altitude of 20,000 m in Earth’s atmosphere: extended incubation periods needed for culture-based assays

On 13 August 2004, an atmospheric sample was collected at an altitude of 20,000 m along a west to east transect over the continental United States by NASA’s Stratospheric and Cosmic Dust Program. This sample was then shipped to the US Geological Survey’s Global Desert Dust program for microbiological analyses. This sample, which was plated on a low nutrient agar to determine if cultivable microorganisms were present, produced 590 small yellow to off-white colonies after approximately 7 weeks of incubation at room-temperature. Of 50 colonies selected for identification using 16S rRNA sequencing, 41 belonged to the family Micrococcaceae, seven to the family Microbacteriaceae, one to the genus Staphylococcus, and one to the genus Brevibacterium. All of the isolates identified were non-spore-forming pigmented bacteria, and their presence in this sample illustrate that it is not unusual to recover viable microbes at extreme altitudes. Additionally, the extended period required to initiate growth demonstrates the need for lengthy incubation periods when analyzing high-altitude samples for cultivable microorganisms.