We anticipate that this document, representing a synthesis of our discussions and supported by literature, will serve as a practical aid to clinicians evaluating patients with suspected encephalitis and will identify key areas and approaches to advance our knowledge of encephalitis.

SUMMARY Background Enterovirus D68 (EV-D68) is implicated in a widespread 2014 outbreak of severe respiratory illness across the United States, and has also been sporadically reported in patients with acute flaccid myelitis (AFM). The association between EV-D68 infection and AFM remains unclear. Methods Here we report metagenomic and molecular epidemiological analyses of 25 AFM cases in California and Colorado from 2012−2014. Findings EV-D68 was detected in respiratory secretions from 7 of 11 (64%) patients comprising two temporally and geographically linked AFM clusters at the height of the 2014 outbreak, and from 12 of 25 (48%) investigated AFM cases overall. Phylogenetic analysis revealed that all AFM-associated EV-D68 sequences grouped into a single novel clade B1 strain that originally emerged in 2010. Out of six observed coding polymorphisms in the clade B1 EV-D68 polyprotein, 5 of 6 polymorphisms were shared between neuropathogenic poliovirus and/or EV-D70. One child with AFM and a sibling with only upper respiratory illness were both infected by identical EV-D68 strains, suggesting a potential role for host-specific factors in differential responses to EV-D68 infection. Notably, EV-D68 viremia was identified in a child experiencing acute neurologic progression of his paralytic illness. Deep metagenomic sequencing of CSF from 14 AFM cases failed to reveal evidence of an alternative infectious etiology to EV-D68. Interpretation Taken together, these findings strengthen the putative association between EV-D68 and AFM, as well as the contention that AFM is a rare yet severe clinical manifestation of EV-D68 infection in susceptible hosts.

Unbiased next-generation sequencing (NGS) approaches enable comprehensive pathogen detection in the clinical microbiology laboratory and have numerous applications for public health surveillance, outbreak investigation, and the diagnosis of infectious diseases. However, practical deployment of the technology is hindered by the bioinformatics challenge of analyzing results accurately and in a clinically relevant timeframe. Here we describe SURPI (''sequence-based ultrarapid pathogen identification''), a computational pipeline for pathogen identification from complex metagenomic NGS data generated from clinical samples, and demonstrate use of the pipeline in the analysis of 237 clinical samples comprising more than 1.1 billion sequences. Deployable on both cloud-based and standalone servers, SURPI leverages two state-of-the-art aligners for accelerated analyses, SNAP and RAPSearch, which are as accurate as existing bioinformatics tools but orders of magnitude faster in performance. In fast mode, SURPI detects viruses and bacteria by scanning data sets of 7-500 million reads in 11 min to 5 h, while in comprehensive mode, all known microorganisms are identified, followed by de novo assembly and protein homology searches for divergent viruses in 50 min to 16 h. SURPI has also directly contributed to real-time microbial diagnosis in acutely ill patients, underscoring its potential key role in the development of unbiased NGS-based clinical assays in infectious diseases that demand rapid turnaround times.

The evolution of virulence was studied in a virus subjected to alternating episodes of vertical and horizontal transmission. Bacteriophage f1 was used as the parasite because it establishes a debilitating but non-fatal infection that can be transmitted vertically (from a host to its progeny) as well as horizontally (infection of new hosts). Horizontal transmission was required of all phage at specific intervals, but was prevented otherwise. Each episode of horizontal transmission was followed by an interval of obligate vertical transmission, followed by an interval of obligate horizontal transmission etc. The duration of vertical transmission was eight times longer per episode in one treatment than in the other, thus varying the relative intensity of selection against virulence while maintaining selection for some level of virus production. Viral lines with the higher enforced rate of infectious transmission evolved higher virulence and higher rates of virus production. These results support the trade-off model for the evolution of virulence.

In the United States, during the past half-century, the number of humans to die of rabies dramatically decreased to an average of 1-2 per year. Although the number of deaths is low, most deaths occur because individuals are unaware that they had been exposed to and infected with rabies virus, and, therefore, they do not seek effective postexposure treatment. Molecular epidemiological studies have linked most of these cryptic rabies exposures to rabies virus variants associated with insectivorous bats. In particular, virus variants associated with 2 relatively reclusive species, the silver-haired bat (Lasionycteris noctivagans) and the eastern pipistrelle (Pipistrellus subflavus), are the unexpected culprits of most cryptic cases of rabies in humans.

Recent phylogenetic analyses of cetacean relationships based on DNA sequence data have challenged the traditional view that baleen whales (Mysticeti) and toothed whales (Odontoceti) are each monophyletic, arguing instead that baleen whales are the sister group of the odontocete family Physeteridae (sperm whales). We reexamined this issue in light of a morphological data set composed of 207 characters and molecular data sets of published 12S, 16S, and cytochrome b mitochondrial DNA sequences. We reach four primary conclusions: (1) Our morphological data set strongly supports the traditional view of odontocete monophyly; (2) the unrooted molecular and morphological trees are very similar, and most of the conflict results from alternative rooting positions; (3) the rooting position of the molecular tree is sensitive to choice of artiodactyls outgroup taxa and the treatment of two small but ambiguously aligned regions of the 12S and 16S sequences, whereas the morphological root is strongly supported; and (4) combined analyses of the morphological and molecular data provide a well-supported phylogenetic estimate consistent with that based on the morphological data alone (and the traditional view of toothed-whale monophyly) but with increased bootstrap support at nearly every node of the tree.

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Rhinovirus is a respiratory virus most typically associated with the common cold and asthma exacerbations, and has not traditionally been considered to play a major role in severe lower respiratory tract infections (LRTIs). As part of a surveillance program for respiratory pathogens of public health importance, children consecutively admitted to intensive care for LRTI at a large tertiary children's hospital were tested with polymerase chain reaction for 11 respiratory viruses and Mycoplasma pneumoniae from February 21 to October 31, 2007; 43 cases were enrolled and rhinovirus was the most frequently detected pathogen, with 21 (49%) positive. Rhinovirus cases frequently were young (median age, 1.4 years [range, 44 days-15 years]), hospitalized for pneumonia (10; 48%), had chronic underlying illnesses (15; 71%), had abnormal chest radiographs (18; 86%), required mechanical ventilation (12; 57%), and had prolonged hospitalization (median length, 7 days [range, 1-29 days]). Coinfection with other viruses or bacteria was common (10; 47%). Rhinovirus may be associated with more severe LRTI in children than previously reported, particularly in the noninfluenza, nonrespiratory syncytial virus season.