No abstract

Viral genome sequencing has guided our understanding of the spread and extent of genetic diversity of SARS-CoV-2 during the COVID-19 pandemic. SARS-CoV-2 viral genomes are usually sequenced from nasopharyngeal swabs of individual patients to track viral spread. Recently, RT-qPCR of municipal wastewater has been used to quantify the abundance of SARS-CoV-2 in several regions globally. However, metatranscriptomic sequencing of wastewater can be used to profile the viral genetic diversity across infected communities. Here, we sequenced RNA directly from sewage collected by municipal utility districts in the San Francisco Bay Area to generate complete and nearly complete SARS-CoV-2 genomes. The major consensus SARS-CoV-2 genotypes detected in the sewage were identical to clinical genomes from the region. Using a pipeline for single nucleotide variant calling in a metagenomic context, we characterized minor SARS-CoV-2 alleles in the wastewater and detected viral genotypes which were also found within clinical genomes throughout California. Observed wastewater variants were more similar to local California patient-derived genotypes than they were to those from other regions within the United States or globally. Additional variants detected in wastewater have only been identified in genomes from patients sampled outside California, indicating that wastewater sequencing can provide evidence for recent introductions of viral lineages before they are detected by local clinical sequencing. These results demonstrate that epidemiological surveillance through wastewater sequencing can aid in tracking exact viral strains in an epidemic context.

Viral genome sequencing has guided our understanding of the spread and extent of genetic diversity of SARS-CoV-2 during the COVID-19 pandemic. SARS-CoV-2 viral genomes are usually sequenced from nasopharyngeal swabs of individual patients to track viral spread. Recently, RT-qPCR of municipal wastewater has been used to quantify the abundance of SARS-CoV-2 in several regions globally. However, metatranscriptomic sequencing of wastewater can be used to profile the viral genetic diversity across infected communities. Here, we sequenced RNA directly from sewage collected by municipal utility districts in the San Francisco Bay Area to generate complete and near-complete SARS-CoV-2 genomes. The major consensus SARS-CoV-2 genotypes detected in the sewage were identical to clinical genomes from the region. Using a pipeline for single nucleotide variant (SNV) calling in a metagenomic context, we characterized minor SARS-CoV-2 alleles in the wastewater and detected viral genotypes which were also found within clinical genomes throughout California. Observed wastewater variants were more similar to local California patient-derived genotypes than they were to those from other regions within the US or globally. Additional variants detected in wastewater have only been identified in genomes from patients sampled outside of CA, indicating that wastewater sequencing can provide evidence for recent introductions of viral lineages before they are detected by local clinical sequencing. These results demonstrate that epidemiological surveillance through wastewater sequencing can aid in tracking exact viral strains in an epidemic context.

Borrelia burgdorferi, the causative agent of Lyme disease, was isolated from the liver of a passerine bird, Catharus fuscescens (veery), and from larval Ixodes dammini (tick) feeding on Pheucticus ludovicianus (rose-breasted grosbeak) and Geothlypis trichas (common yellowthroat). In indirect immunofluorescence antibody tests, isolates reacted with polyclonal and monoclonal (H5332) antibodies. Studies on the DNA composition of the veery liver isolate and the strain cultured from an I. dammini larva indicated that both were B. burgdorferi and not Borrelia anserina or Borrelia hermsii. The veery liver isolate infected hamsters and a chick. In contrast, B. anserina infected chicks but not hamsters. B. burgdorferi is unique among Borrelia spp. in being infectious to both mammals and birds. We suggest that the cosmopolitan distribution of B. burgdorferi may be caused by long-distance dispersal of infected birds that serve as hosts for ticks. In the northeastern United States, Borrelia burgdorferi (21) is transmitted primarily by Ixodes dammini (15), a tick that parasitizes a variety of mammals and birds (3, 5, 18, 30, 33). This spirochete has been isolated, cultured, and identified from I. dammini (4-6, 15, 22, 25), Ixodes pacificus (17),

Genetic studies were performed on the following spirochetes: three Lyme disease spirochetes isolated from Ixodes ticks and from human spinal fluid; three species of North American borreliae; four species of Treponema; and two species of Leptospira. The mol% G+C values for Lyme disease spirochetes were 27.3 to 30.5%, similar to values of 28.0 to 30.5% for Borrelia species but different from the values for Leptospira or Treponema species which ranged from 35.3 to 53%. Lyme disease spirochetes represent a new species of Borrelia, with DNA homologies of 31 to 59% with the three North American strains of Borrelia studied. These studies also showed that Lyme disease spirochetes from three sources constituted a single species, with DNA homologies ranging from 76 to 100%. A high degree of relatedness was also seen between the three North American borreliae, with homology varying from 77 to 95%, indicating that these spirochetes represent a single species. Lyme disease spirochetes and Borrelia species exhibited almost no homology with Leptospira and Treponema species (0 to 2%). Plasmids were detected in the three Lyme disease spirochetes and in the three North American borreliae. Lyme disease was first recognized in 1975 (2, 21) and occurs in the spring, summer, and fall, beginning with a red skin lesion, erythema chronicum migrans (ECM) (2, 21). Accompanying this are headache, myalgia, malaise, and arthralgias, which last for several weeks. Weeks or months after the early clinical phase of the disease, patients may develop neurological or cardiac abnormalities (21). Patients may go on to develop intermittent attacks of arthritis which may become chronic, resulting in destruction of bone and cartilage and in the large joints (21). Epidemiological studies suggested the involvement of an unknown agent transmitted by the deer tick, Ixodes dammini. Penicillin improved or eliminated the symptoms, suggesting involvement of a penicillin-sensitive bacterium (21). The organism responsible for the disease was discovered during a tick and pathogen survey on Shelter Island in New York. Sixty-one percent of the I. dammini ticks examined contained spirochetes that were antigenically reactive with sera from patients in convalescent stages of Lyme disease (6). The same spirochete was isolated from blood, cerebrospinal fluid, and skin lesions of patients with Lyme disease, confirming the role of these spirochetes in this malady (2, 21). Clinical and epidemiological similarities between Lyme disease and ECM in Europe led Barbour et al. to study Ixodes ricinus, the tick incriminated as the vector of ECM in Switzerland. Thirty six percent of the I. ricinus ticks examined contained spirochetes that were similar, if not identical, to the New York-isolated Lyme disease spirochetes (1). Lyme disease in the United States covers three geographical areas: the Northeast and the Midwest, where I. dammini carries the spirochetes, and the west, with Ixodes pacificus ticks serving as the vector. Isolated cases of Lyme disease have been reported in Texas...