The lion's share of bacteria in various environments cannot be cloned in the laboratory and thus cannot be sequenced using existing technologies. A major goal of single-cell genomics is to complement gene-centric metagenomic data with whole-genome assemblies of uncultivated organisms. Assembly of single-cell data is challenging because of highly non-uniform read coverage as well as elevated levels of sequencing errors and chimeric reads. We describe SPAdes, a new assembler for both single-cell and standard (multicell) assembly, and demonstrate that it improves on the recently released E+V-SC assembler (specialized for single-cell data) and on popular assemblers Velvet and SoapDeNovo (for multicell data). SPAdes generates single-cell assemblies, providing information about genomes of uncultivatable bacteria that vastly exceeds what may be obtained via traditional metagenomics studies. SPAdes is available online ( http://bioinf.spbau.ru/spades ). It is distributed as open source software.

SPAdes, a new genome assembly algorithm and its applications to single-cell sequencing ( 7th Annual SFAF Meeting, 2012)

https://www.thelancet.com/pdfs/journals/lancet/PIIS0140-6736(07)61779-6.pdf

Infection with chikungunya virus in Italy: an outbreak in a temperate region

Background: Dengue is a growing problem both in its geographical spread and in its intensity, and yet current global distribution remains highly uncertain. Challenges in diagnosis and diagnostic methods as well as highly variable national health systems mean no single data source can reliably estimate the distribution of this disease. As such, there is a lack of agreement on national dengue status among international health organisations. Here we bring together all available information on dengue occurrence using a novel approach to produce an evidence consensus map of the disease range that highlights nations with an uncertain dengue status. Methods/Principal Findings: A baseline methodology was used to assess a range of evidence for each country. In regions where dengue status was uncertain, additional evidence types were included to either clarify dengue status or confirm that it is unknown at this time. An algorithm was developed that assesses evidence quality and consistency, giving each country an evidence consensus score. Using this approach, we were able to generate a contemporary global map of national-level dengue status that assigns a relative measure of certainty and identifies gaps in the available evidence. Conclusion: The map produced here provides a list of 128 countries for which there is good evidence of dengue occurrence, including 36 countries that have previously been classified as dengue-free by the World Health Organization and/or the US Centers for Disease Control. It also identifies disease surveillance needs, which we list in full. The disease extents and limits determined here using evidence consensus, marks the beginning of a five-year study to advance the mapping of dengue virus transmission and disease risk. Completion of this first step has allowed us to produce a preliminary estimate of population at risk with an upper bound of 3.97 billion people. This figure will be refined in future work.

/pdf/refining-the-global-spatial-limits-of-dengue-virus-f8emry0wk5.pdf

Refining the global spatial limits of dengue virus transmission by evidence-based consensus

Background
Accurate rapid diagnostic tests for SARS‐CoV‐2 infection could contribute to clinical and public health strategies to manage the COVID‐19 pandemic. Point‐of‐care antigen and molecular tests to detect current infection could increase access to testing and early confirmation of cases, and expediate clinical and public health management decisions that may reduce transmission.

Objectives
To assess the diagnostic accuracy of point‐of‐care antigen and molecular‐based tests for diagnosis of SARS‐CoV‐2 infection. We consider accuracy separately in symptomatic and asymptomatic population groups.

Search methods
Electronic searches of the Cochrane COVID‐19 Study Register and the COVID‐19 Living Evidence Database from the University of Bern (which includes daily updates from PubMed and Embase and preprints from medRxiv and bioRxiv) were undertaken on 30 Sept 2020. We checked repositories of COVID‐19 publications and included independent evaluations from national reference laboratories, the Foundation for Innovative New Diagnostics and the Diagnostics Global Health website to 16 Nov 2020. We did not apply language restrictions.

Selection criteria
We included studies of people with either suspected SARS‐CoV‐2 infection, known SARS‐CoV‐2 infection or known absence of infection, or those who were being screened for infection. We included test accuracy studies of any design that evaluated commercially produced, rapid antigen or molecular tests suitable for a point‐of‐care setting (minimal equipment, sample preparation, and biosafety requirements, with results within two hours of sample collection). We included all reference standards that define the presence or absence of SARS‐CoV‐2 (including reverse transcription polymerase chain reaction (RT‐PCR) tests and established diagnostic criteria).

Data collection and analysis
Studies were screened independently in duplicate with disagreements resolved by discussion with a third author. Study characteristics were extracted by one author and checked by a second; extraction of study results and assessments of risk of bias and applicability (made using the QUADAS‐2 tool) were undertaken independently in duplicate. We present sensitivity and specificity with 95% confidence intervals (CIs) for each test and pooled data using the bivariate model separately for antigen and molecular‐based tests. We tabulated results by test manufacturer and compliance with manufacturer instructions for use and according to symptom status.

Main results
Seventy‐eight study cohorts were included (described in 64 study reports, including 20 pre‐prints), reporting results for 24,087 samples (7,415 with confirmed SARS‐CoV‐2). Studies were mainly from Europe (n = 39) or North America (n = 20), and evaluated 16 antigen and five molecular assays.

We considered risk of bias to be high in 29 (37%) studies because of participant selection; in 66 (85%) because of weaknesses in the reference standard for absence of infection; and in 29 (37%) for participant flow and timing. Studies of antigen tests were of a higher methodological quality compared to studies of molecular tests, particularly regarding the risk of bias for participant selection and the index test. Characteristics of participants in 35 (45%) studies differed from those in whom the test was intended to be used and the delivery of the index test in 39 (50%) studies differed from the way in which the test was intended to be used. Nearly all studies (97%) defined the presence or absence of SARS‐CoV‐2 based on a single RT‐PCR result, and none included participants meeting case definitions for probable COVID‐19.

Antigen tests

Forty‐eight studies reported 58 evaluations of antigen tests. Estimates of sensitivity varied considerably between studies. There were differences between symptomatic (72.0%, 95% CI 63.7% to 79.0%; 37 evaluations; 15530 samples, 4410 cases) and asymptomatic participants (58.1%, 95% CI 40.2% to 74.1%; 12 evaluations; 1581 samples, 295 cases). Average sensitivity was higher in the first week after symptom onset (78.3%, 95% CI 71.1% to 84.1%; 26 evaluations; 5769 samples, 2320 cases) than in the second week of symptoms (51.0%, 95% CI 40.8% to 61.0%; 22 evaluations; 935 samples, 692 cases). Sensitivity was high in those with cycle threshold (Ct) values on PCR ≤25 (94.5%, 95% CI 91.0% to 96.7%; 36 evaluations; 2613 cases) compared to those with Ct values >25 (40.7%, 95% CI 31.8% to 50.3%; 36 evaluations; 2632 cases). Sensitivity varied between brands. Using data from instructions for use (IFU) compliant evaluations in symptomatic participants, summary sensitivities ranged from 34.1% (95% CI 29.7% to 38.8%; Coris Bioconcept) to 88.1% (95% CI 84.2% to 91.1%; SD Biosensor STANDARD Q). Average specificities were high in symptomatic and asymptomatic participants, and for most brands (overall summary specificity 99.6%, 95% CI 99.0% to 99.8%).

At 5% prevalence using data for the most sensitive assays in symptomatic people (SD Biosensor STANDARD Q and Abbott Panbio), positive predictive values (PPVs) of 84% to 90% mean that between 1 in 10 and 1 in 6 positive results will be a false positive, and between 1 in 4 and 1 in 8 cases will be missed. At 0.5% prevalence applying the same tests in asymptomatic people would result in PPVs of 11% to 28% meaning that between 7 in 10 and 9 in 10 positive results will be false positives, and between 1 in 2 and 1 in 3 cases will be missed.

No studies assessed the accuracy of repeated lateral flow testing or self‐testing.

Rapid molecular assays

Thirty studies reported 33 evaluations of five different rapid molecular tests. Sensitivities varied according to test brand. Most of the data relate to the ID NOW and Xpert Xpress assays. Using data from evaluations following the manufacturer’s instructions for use, the average sensitivity of ID NOW was 73.0% (95% CI 66.8% to 78.4%) and average specificity 99.7% (95% CI 98.7% to 99.9%; 4 evaluations; 812 samples, 222 cases). For Xpert Xpress, the average sensitivity was 100% (95% CI 88.1% to 100%) and average specificity 97.2% (95% CI 89.4% to 99.3%; 2 evaluations; 100 samples, 29 cases). Insufficient data were available to investigate the effect of symptom status or time after symptom onset.

Authors' conclusions
Antigen tests vary in sensitivity. In people with signs and symptoms of COVID‐19, sensitivities are highest in the first week of illness when viral loads are higher. The assays shown to meet appropriate criteria, such as WHO's priority target product profiles for COVID‐19 diagnostics (‘acceptable’ sensitivity ≥ 80% and specificity ≥ 97%), can be considered as a replacement for laboratory‐based RT‐PCR when immediate decisions about patient care must be made, or where RT‐PCR cannot be delivered in a timely manner. Positive predictive values suggest that confirmatory testing of those with positive results may be considered in low prevalence settings. Due to the variable sensitivity of antigen tests, people who test negative may still be infected.

Evidence for testing in asymptomatic cohorts was limited. Test accuracy studies cannot adequately assess the ability of antigen tests to differentiate those who are infectious and require isolation from those who pose no risk, as there is no reference standard for infectiousness. A small number of molecular tests showed high accuracy and may be suitable alternatives to RT‐PCR. However, further evaluations of the tests in settings as they are intended to be used are required to fully establish performance in practice.

Several important studies in asymptomatic individuals have been reported since the close of our search and will be incorporated at the next update of this review. Comparative studies of antigen tests in their intended use settings and according to test operator (including self‐testing) are required.

/pdf/rapid-point-of-care-antigen-and-molecular-based-tests-for-2a2qjx1fq9.pdf

Rapid, point-of-care antigen and molecular-based tests for diagnosis of SARS-CoV-2 infection.

The first edition of this reference work was published in 1898, and the last update was published in 1972. This current edition is written by six major contributors and contains either rewritten or new chapters, including one 29-page chapter entitled "Ophthalmology in the Tropics" by F. C. Rodger, MD. Not only is this material valuable to physicians in endemic areas, but it is also important for travelers to the tropics who may return home with these diseases. Most of the chapters discuss the following aspects of tropical disease: cause, transmission, immunology, epidemiology, geographical distribution, pathologic condition, clinical findings, and diagnosis (including laboratory findings, treatment, and prevention). Beside chapters on infections, there are chapters on disorders due to heat, nutritional diseases, and venoms and poisons, and appendices on protozoology, helminthology, entomology, and clinical pathologic conditions. Excellent illustrations of end-stage pathologic conditions are disconcerting. Ophthalmologists would be most interested in the discussion

Manson's Tropical Diseases

Background. An outbreak of chikungunya virus infection emerged in the southwest Indian Ocean islands in 2005, spread out to India, and resulted in an ongoing outbreak that has involved >1.5 million patients, including travelers who have visited these areas.Methods. Our study investigated 69 travelers who developed signs and symptoms compatible with chikungunya fever after returning home from countries involved in the epidemic. Twenty cases of infection that were confirmed by serological analysis, polymerase chain reaction, and/or cell culture were investigated.Results. All patients experienced flulike symptoms with fever and joint pain. No serious complications were observed, but 69% of the patients had persistent arthralgia for >2 months, and 13% had it for >6 months. Viral RNA could be detected in blood samples using reverse-transcriptase polymerase chain reaction in 4 of 4 patients who presented to a health care facility during their first week of illness, and the virus was successfully isolated from blood samples obtained from 2 of these patients. Chikungunya virus–specific immunoglobulin M and/or immunoglobulin G antibodies were detected in all patients. However, initial testing of serum samples yielded negative results for 3 of 5 patients during the first week.Conclusions. Chikungunya fever must be considered in travelers who develop fever and arthritis after traveling to areas affected by an ongoing epidemic. Related arthritis mainly affects smaller joints and often persists for extended periods. Serological testing may have negative results during the first week of the disease; diagnosis using polymerase chain reaction appears to be more reliable during this time. Travelers to areas of epidemicity should be informed of the risk of infection and of adequate preventive measures, such as protection against mosquitos.

/pdf/chikungunya-fever-in-travelers-clinical-presentation-and-4gyw3sw4o1.pdf

Chikungunya Fever in Travelers: Clinical Presentation and Course

Tick-borne encephalitis: Pathogenesis and clinical implications

Background: The current outbreak of SARS-CoV-2 has spread to almost every country with more than three million confirmed cases and over two hundred thousand deaths as of April 28, 2020. Rapid first-line testing protocols are needed for outbreak control and surveillance. 

Methods: We used computational and manual design to generate a suitable set of reverse transcription recombinase polymerase amplification (RT-RPA) primer and exonuclease probe, internally quenched (exo-IQ) probe sequences targeting the SARS-CoV-2 N gene. RT-RPA sensitivity was determined by amplification of in vitro transcribed RNA standards. Assay selectivity was demonstrated with a selectivity panel of 32 nucleic acid samples derived from common respiratory viruses. To validate the assay against full-length SARS-CoV-2 RNA, total viral RNA derived from cell culture supernatant and 19 nasopharyngeal swab samples (8 positive and 11 negative for SARS-CoV-2) were screened. All results were compared to established RT-qPCR assays. 

Results: The 95% detection probability of the RT-RPA assay was determined to be 7.74 (95% CI: 2.87 - 27.39) RNA copies per reaction. The assay showed no cross-reactivity to any other screened coronaviruses or respiratory viruses of clinical significance. The developed RT-RPA assay produced 100% diagnostic sensitivity and specificity when compared to RT-qPCR (n=20). 

Conclusion: With a run time of 15 to 20 minutes and first results being available in under 7 minutes for high RNA concentrations, the reported assay constitutes one of the fastest nucleic acid based detection methods for SARS-CoV-2 to date and may provide a simple to use alternative to RT-qPCR for first-line screening at the point of need.

/pdf/rapid-detection-of-sars-cov-2-by-low-volume-real-time-single-2vzbg33a42.pdf

Rapid Detection of SARS-CoV-2 by Low Volume Real-Time Single Tube Reverse Transcription Recombinase Polymerase Amplification Using an Exo Probe with an Internally Linked Quencher (Exo-IQ).

Tick-borne encephalitis (TBE) is the main tick-borne virus infection in Eurasia. It is prevalent across the entire continent from Japan to France and occurs in endemic foci. Expansion of prevalence in areas including northern Russia, Sweden, and Finland has been observed in recent years. Ticks are the most important vectors and may transmit the TBE virus to animals and humans. TBE can also be transmitted to humans in milk containing the virus. TBE has been implicated as a travel-acquired illness and there are isolated reports of its occurrence in countries outside the known areas of prevalence. Therefore, TBE should be included in the differential diagnosis for all central nervous system diseases inside or outside endemic areas.

Epidemiology and distribution of tick-borne encephalitis

Historically, flea-borne diseases are among the most important medical diseases of humans. Plague and murine typhus are known for centuries while the last years brought some new flea-transmitted pathogens, like R. felis and Bartonella henselae. Dogs may play an essential or an accidental role in the natural transmission cycle of flea-borne pathogens. They support the growth of some of the pathogens or they serve as transport vehicles for infected fleas between their natural reservoirs and humans. More than 15 different flea species have been described in domestic dogs thus far. Several other species have been found to be associated with wild canids. Fleas found on dogs originate from rodents, birds, insectivores and from other Carnivora. Dogs therefore may serve as ideal bridging hosts for the introduction of flea-borne diseases from nature to home. In addition to their role as ectoparasites they cause nuisance for humans and animals and may be the cause for severe allergic reactions.

/pdf/fleas-as-parasites-of-the-family-canidae-4sj1iqb0ze.pdf

Gerhard Dobler

Papers

Chikungunya Fever in Travelers: Clinical Presentation and Course

Tick-borne encephalitis: Pathogenesis and clinical implications

Rapid Detection of SARS-CoV-2 by Low Volume Real-Time Single Tube Reverse Transcription Recombinase Polymerase Amplification Using an Exo Probe with an Internally Linked Quencher (Exo-IQ).

Epidemiology and distribution of tick-borne encephalitis

Fleas as parasites of the family Canidae