SARS-CoV-2 sequencing reveals rapid transmission from college student clusters resulting in morbidity and deaths in vulnerable populations

College reop ening decisions during t he S ARS-CoV-2 pandemic repres ent a t rad e-off between competing risks to studen ts, faculty and staff, and c ollege finances. Additio nally, risks taken i n reopeni ng colleges can impose significant burdens on indivi duals living in surrounding communities. Many colleges tha t reope ned for in-pers on instruc tion have repor ted fre quen t SARS-CoV-2 outbre aks. La Crosse County, Wisconsin exp erienc ed a substa ntial S AR S-CoV-2 outbreak (2,002 cases in Septem ber 2020) that coincided with th e ret urn t o in-person in struction a t th ree l ocal academic insti tuti ons. Ge nomic sequencing of SARS-CoV-2 cases in La Cr osse during tha t peri od found rapid expa nsion of two viral substrains. Altho ugh the majori ty of cases were among college-age individuals, fr om a total of 111 genomes sequenc ed we iden tified rapid transmission of the virus in to more vulne rable popul atio ns. Eight sampled genomes r epres ent ed two independ ent t ransmission even ts into t wo skilled nursing facilities, resul ting in two fatali ties . Our s tudy highlights the very significant risks imposed by college administra tor r eope ning decisions, no t just on college-associa ted popul atio ns, but on vulnerable individuals in surrounding communiti es. . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint NOTE: This preprint reports new research that has not been certified by peer review and should not be used to guide clinical practice. 2

The SARS-CoV-2 betacoron avirus is a novel respir ato ry pathogen which eme rged i n Wuhan, China in lat e 2019 (1). Numerous teams aro und th e world have produc ed a rapi dly increasing number of publicly available sequ ences (116,650 in GISA ID on 9/28/20 ) with obvious sequence divers ification as viral replicati on and spr ead has conti nued . These seque nce varian ts, inher ited in each cycle of viral replicati on, allow th e gene rati on of phylogenetic t re es thro ugh which the evoluti on of the virus can be traced . Individual viruses acq uire mut ati ons during replica tion a t an appr oxima te rate of 1-2 sites pe r month (2). This mutation ra te pr ovides good resolu tion for bo th global and local t racking of viral substrains. Because viral replica tion occu rs in individuals, tracking th ese inhe rit ed variants acts as a proxy for tr acking the spre ad of the virus through popul ations of individuals. Star ting in late M arch 2020, we have mo nitor ed the in trod uction and sp read of S ARS-CoV-2 to the service are a of the Gunders en Heal th System, an in tegra ted h ealthca re system he adqu ar ter ed in La Crosse, WI and p roviding care in 21 coun ties in southwes ter n Wisconsin, no rth eas tern I owa and southeas te rn Minneso ta . We have p erfo rmed whole viral genome s equencing on 514 positive cases from this region, spanni ng thre e sta tes w hich have adopt ed mitigati on appr oache s of differing intensity. In Sept ember 2020, La Crosse County (W isconsin) experi enced a ra pid increas e in cases, strongly driven by infections among the 10-19 and 20-29 age groups. The pe riod coincided with t he ret urn of college studen ts to th ree ins titu tions of higher e ducation in th e city of La Crosse: Universi ty of Wisconsin La Crosse, West ern Technical Coll ege and Viterbo University, with tot al enrollm ents of 10,558, 4,004 and 2,592, respec tively (3). Our regional SA R S-CoV-2 surveillance and sequencing pro gram capture d many of these gen omes, allowing assessmen t of t he substrain con tribu tions t o the o utbr e ak and analysis of the speed with which vulnerabl e non-stud en t popula tions were affect ed.

Specimens Specimens were nas opharynge al swabs taken for diagnostic purp oses and evalu at ed for SARS-CoV-2 positivity at th e Gund erse n Medical Fou ndation ’s Molecula r Diagnostics Labora t ory using the CDC 2019- nCoV Real-Time RT-PCR Diagnostic assay. This study includes positive cases diagno sed betwe en . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 3 3/18/2020 and 9/23 /2020. Ethical appro val was obtained from th e Gund ersen H e alth System Institution al Review Boar d (#2-20-03-008 ; PI: Kenny) to perform addi tional n ex t-genera tion se qu encing on remnan t specimens afte r completi on of diagnostic testi ng. Samples for th is study inclu de only those testing posi tive in Gund ersen H ealt hcare System’s diagnostic labo rato ries and , as such, do not include other cas es from the r egion which may have been t est ed in oth er heal thcar e syst ems and/or public health la bora tori es. RNA isolation and cDNA synthesis Nasopha ryngeal swabs were immers ed in liquid medium. RN A was isolated using QIAmp Viral RN A Mini kit (Qiagen, Ge rmantown, MD). cDNA was synthesized from 5 μl of purified RNA u sing ProtoScript I I First Strand cDNA Synt hesis Kit (New England Biolabs, Ipswich, M A) with random he xa mers. Next Generation sequencing We used th e Ion AmpliSeq SA RS-CoV-2 P anel (Thermo-Fisher, Walt ham, MA) to s equence 237 viral specific targets enc ompassing the compl ete viral gen ome from cDNA derived fro m SARS-COV-2-positive clinical specimens. Barc oded multipl ex ed libraries wer e prep are d in batches of eig ht on the Ion Chef (Thermo-Fisher) and sequenc ed on th e I on Torren t S5 (Thermo-Fisher). Bioinformatics Demultiplex ed Tmap-aligned bam files were pr oduced by the Ion Torr ent S5 . To overcome difficulties caused by read soft-clipping by this non- splice aware aligne r, individual sequ ence reads were ex tract ed using samtools (4) and re-aligned to th e SARS-CoV-2 genome using a splice-aware aligner, hisa t2 (5). These bam files were subject ed to QC re view in IGV. Samples passing QC had a consensus sequence computed accordi ng to th e metho d of Cavener (6). Consensus sequences wer e re porte d to G ISA ID (7). For phylogene tic inferenc e (i.e. t o det er mine the hie rarchy of case rel ationshi ps) samples were integra ted with associ ate d metad ata and aligned on a local implemen ta tion of Ne xtSt rain (8) using augur and displayed via a web browse r u sing auspice. Chorople th maps were ge n erat ed using folium.

The cumulative histo ry of SARS-CoV-2 ca ses, stra tified by age, for La Crosse count y is shown in Figure 1. Notab le even ts along this tim eline includ e the r eope ning of bars in Wisconsin (5/14/2020) and the onset . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 4 of warm summer weather (first day abov e 80’F was on Memorial Day We ekend o n 5/24/2020 ) which preced ed a rapid ris e in cases in ea rly-mid June. Case grow th continu ed at a slower ra te for much of the summer before ab ruptly incr easing in th e second week of Sept ember , coinciding with the r etur n of studen ts to th ree coll eges locat ed in th e city of La Crosse. Analysis of cases by census tract showed that the most r apid case growth was occurri n g the immediat e vicinity of the thr ee coll eges, an ar ea of high density stude nt housing (Figure 2). Betwe en 3/18/20 and 9/23/20 we obtain ed 514 high quality genomes (Figure 3A) from surveillance in a 21-county region of western Wisconsin, southeas te rn Minneso ta and n orth east e rn Iowa. Clades a re colored for e ase of visualizatio n and incl ude several epidemiologically r elevant o utbre aks. Significant examples includ e (i) an outbre ak originat ing in a meatpacking plant in Postville , IA (9) and (ii) a large outbr eak associat ed with a clust er of bar s in downtown La Crosse in June/July. Fig ure 3B highlights newly sequenced genom es in our region in Septemb er 2020. The ove rwhelming majority of case growth in La Crosse county was explaine d by two clusters, d esignate d “College A” and “C ollege B”. The small light-blue cluste r at th e 11 o’clock positi on repr esen ts a substrain d et ected in the city of Holmen, which did not involve college stud ents , while th e othe r highlighted clad es rep resen ted c ase growth in our region outsid e of La Crosse county. He re we focus on the analysis of the College A and College B clusters . During this period, al l thr ee acad emic institu tions post ed “COVID19 dashboards” on their websi tes, analysis of which indicated tha t the majo rity of cases were associa ted with t he lar gest institution (University of Wisconsin – La Crosse, “UWL”), followed by Viterbo University and with relatively few cases affecting studen ts enroll ed at West ern Technical College . UWL ran its own s urveillance and diagnostic program for st uden ts living in dormitori es (appro ximat ely 3000 studen ts), but did not d eploy resources sufficient to provide a ny surveillance for its large off-campus studen t po pulation . Our sequencing program cap tur ed all positiv e tests cond ucted a t th e Gund ersen H eal th System molecular diagnostic labor ato ry, a service availed o f by many members of the off-campus populatio n. Accordi ngly, while we generally lack genomes from th e studen t dorm popul ations , our conside rable cover age of the off-campus population and th e presuma bly large amount of int ermixing th at occ urred b etwee n the on and-off campus student popul ations du e to the p revalen t “party cultu re” (10,11) makes it unlikely that a substanti al outb reak comprising an unr el ated subs train might have occur red in an d been res trict ed to the dorm-dwelling popul ation . . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 5 Demographic detai ls of the 111 individua ls carrying the College A and B subs trains are shown in Table 1. In tot al, 40.5% of the sp ecimens were ob tained from individuals age d betwe en 17 and 24, with a furthe r 16.2% from the 25-29 age group. For the entir e cohort , 57.7% of cases affected fe males, alth ough cases were more evenly distributed be tween s exes among t he 17-29 year old grou p . 19 cases (17.1%) af fected individuals aged over 60, including 4 case s among individuals in their 90s. This pat tern of spre ad from clusters of younger age individuals in to more vulnerabl e age groups is consist ent with observations of a time-lag in which cases among older indi viduals tend to rise afte r outb reaks amo ng young individuals (12). Although this sta tistical t rend is evi dent from a na tionwide an alyses (12), our ability to es tablish direct gen etic links using sequencing be t ween the two age groups provid es compelling evidence th at risks of rapid spread of SARS-CoV-2 among college-age individuals are no t limit ed to college enviro ns but pose a direc t thr eat to old er pers ons in t he surroundi ng community. Table 1: Demographics of individuals associated with College A and College B clusters College A (n=68) College B (n=43) Both Strains (n=111) Age Group 0-5 0 (0% ) 1 (2.3%) 1 (0.9%) 6-16 1 (1.5%) 1 (2.3%) 2 (1.8%) 17-24 24 (35.3%) 21 (48.8%) 45 (40.5%) 25-29 15 (22.1%) 3 (7% ) 18 (16.2%) 30-39 6 (8.8%) 4 (9.3%) 10 (9.0%) 40-49 9 (13.2%) 1 (2.3%) 10 (9.0%) 50-59 4 (5.9%) 2 (4.7%) 6 (5.4%) 60-69 4 (5.9%) 3 (7% ) 7 (6.3%) 70-79 3 (4.4%) 1 (2.3%) 4 (3.6%) 80-89 2 (4.7%) 2 (4.7%) 4 (3.6%) 90-99 0 (0% ) 4 (9.3%) 4 (3.6%) Sex (All ages) Male 30 (44.1%) 17 (39.5%) 47 (42.3%) Female 38 (55/9 %) 26 (60.5%) 64 (57.7%) Sex (Ages 17-29) Male 22 (56.4%) 10 (41.6%) 32 (50.8%) Female 17 (43.6%) 14 (58.3%) 31 (49.2%) . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 6 Detailed phylogen etic t re es for both Coll ege A and B clust ers ar e shown in Figure 4. Trees ar e scaled t o show genetic divergence of t he virus, suc h that sampl es aligned ver tically are gen etically iden tical and placement of a sample one incr ement to the right ind icates acquisition of a single new mu tation . Wi thin both cluste rs, we found evidence of subg roups sharing iden tical viruses which typi cally reflect individuals infected in quick succession from a common source (e.g. at o ne or mor e parti es). Gr oups of cases containing five or more gen etical ly identical gen omes are highligh ted in or a nge, and th e median age of individuals in these four grou ps were 20.5, 21, 25 an d 29. This patt ern is co nsistent with rapid spread of SARS-CoV-2 among the younge r age group. Al though bo th College A an d B clusters included older individuals (See Tabl e 1), analysis of the College B cluste r indicat ed tha t this particula r substr ain had been transmit ted in to two skilled nursing facilities. The sequencing data reve aled indep enden t clusters affecting th ree p atie nts in one fa cility and two staff and five patien ts in th e second facility. Two of these pa tien ts died from COVID-19.

This study provides an inte rim yet compr ehensive look a t a still-ongoing outb reak cente red on t he neighborho ods surroun ding college cam puses in a midwestern ci ty, La Crosse, WI . While t he 111 genomes sampled r epres ent a frac tion of the to tal cases in La Crosse coun ty in thi s Septembe r outb reak, the da ta app ear sufficient t o draw some importan t conclusions abou t th e natu re of the outb reak and early consequ ences for vulnerabl e popul ations, as well as pr ompting conside rati o n of whether various mitigation st rat egies adop ted elsewher e could have been h elpful. Firstly, this larg e outb reak was overwhel mingly driven by just two substrains of SARS-CoV-2 which was quite surp rising. Res tar ting of in-person educatio n involved the r etu rn of many thousands of studen ts from a wide range of cities and small tow ns throughou t the M idwest a t a time wh en SARS-CoV-2 cases in Midwester n 18-22 year olds were incr easing at a ra te mor e th an twice th e nati onal average for tha t age group (13). Accordingly, we had exp ected gre at er viral diversi ty consistent wi th multiple independ ent viral in troduc tions. In contr ast, th e major signal det ecte d was for just two substra ins which spread very rapidly among an a t-risk pop ulation which socialize d rep eat edly at hi gh-density alcohol- fueled indoor and outd oor ga therings . W e cannot e xclude the possibili ty tha t the r e were addi tional contempor aneo us viral intro ductions an d that t hese two substrains for tuit ously happene d to be the earlies t to b e exp osed t o large suscep tibl e popula tions at events favoring rapid S A RS-CoV-2 spread such that o the r pot ential subs trains did n ot h ave the opp ortun ity to access such a larg e susceptible . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 7 populati on. N evert heless, o ur da ta imply that th e overwhelming majori ty of stude nts were SAR S-CoV-2- negative upon r etu rn to the campus envi rons and tha t rapi d local spre ad, ra the r t han import ed cases, was the primary cause of th e local out br eak. Natio nally, several unive rsiti es made ver y considerable financial inves tments in p re-tes ting all stude nts before th eir r etu rn to campus, a mitiga ti on stra tegy that was no t at temp ted in La Crosse. Our da ta suggest that , had th at st rat egy been emp loyed in La Crosse, it would have had a n egligible impact on preventing the outbre ak. A t best, i t might have delayed a n inevitabl e ou tcom e by a few days. Clearly, introduc tion of just two viral substr ains was sufficient to fuel this very significant outbr eak. In a s et ting where stud ents from multipl e colleges in terac t with each o the r and with ot her r e sidents of a city with pre-exis ting SARS-COV-2 cases, studen t b ehavior will likely lead to significant ou tb reaks wheth er th e initial stud ent-impor ted viral inp ut is larg e or small. Testing of symptomatic and asymptoma t ic students aft er college opening is likely a more useful interven tion, if pe rformed a t a sufficient frequency. Wh en modeling multipl e scen arios to mitiga te SARS-CoV-2 spread during college r e-ope ning for an 80-day fall semester, Palti el a nd colleagues estimat ed tha t t esting th e enti re stu den t population every two days could limit cu mulative infections to 4.9% of students, while we ekly testing would result in 37% of studen ts becoming infected (14). Import antly, bo th estima tes involved ass umptions of strong adh erenc e to mask-wearing, handwashi ng, social-distancing, de-densifying classroo ms and limiting bathro om sharing. The U WL plan for its 10,500 studen t body involved asymptomatic t es ting of its dormitory r esiden t popula tion (3000 students) once every 14 days with no surveillance t estin g among the much larger off-campus population . Considering that Palti el estim ate d tha t tes ting 100% of an idealized guid eline-adh eren t stud e nt popula tion weekly would still lead to a cumulative infecti on rate of 37%, it se ems likely that t esting o nly 28% of an actual studen t popula tion a t half tha t frequenc y would be utterly inad equa te t o manag e inevitabl e outb reaks. Success in Paltiel’s model also r equir ed e fficiently functioning quaranti ne and isol ation do rmitori es, a situatio n at varianc e with th e chaotic sit uation a t UWL where s tuden t rep ort ers described individuals quaran tined while the r esults of thei r tes ts were pend ing, par tying in the quar anti ne dorm with COVID19-positive individuals, and the n being releas ed when th eir pr e-quara ntin e test r etu rned n egative (15). . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 8 We de tect ed th ese stud ent-amplified inf ection clust ers as par t of a 21-county surveillance program in the Gu nders en Heal thcar e System’s servi ce area . Becaus e this progr am was not st udent focused, we were collecti ng and sequencing o ther co mmunity cases in parallel . This allowed u s to quickly detect the overspill from the colleg e-age popula tion in older adul ts. Whil e thes e findings were consisten t with public health exp ecta tions abou t risk to older popul ation in s ett ings of wide community spread and with epidemiological st udies showing a sta tist ical association in which case incre ases i n young adults are typically followed by cases among older adults (12), our ability to gen etically link these grou ps of cases provides direc t evidence of t ransmission betwee n thes e differen t age groups. Of particula r concern, was the rapi d tra nsmission of one of these S A RS-CoV-2 substrains into two skilled nurs ing facilities, causing sustained ou tbr eaks with two fatali ties s o far. Our first cas e of what we came to c all the “College B” cluster was collect ed on 8/27/20, a time when cases were slowly but ste adily risin g (Figure 1). The inflection poin t on the curve when cases began to ris e more r apidly was on 9/10/ 20 (Figu re 1) and the first skilled nursing facility patien t speci men associate d with this cluste r was det ected on 9/14/20.

For college clust ers, much concern h as focused on higher risks to old er faculty/sta ff as well as less frequen t but significant advers e outcom es in younger individuals. In a dditio n, uni versity leade rs clearly face a range of concerns including very significant financial pressur es. Given the d escribed t esting strat egy, the highly contagio us prope rti e s of SARS-CoV-2, news reports of widespread ou tbre aks in other coll ege communiti es tha t had at te mpted re turn to in-perso n instruc tion at earlie r dat es (16,17) and the well-und ersto od risk-embracing natur e of college stud ents (18), the out b reak tha t occurr ed in La Crosse in Septemb er 2020 was likely inevitable o nce th e decision to ret urn to i n-person instr uction was taken. Our stu dy highlights th e very significant risks imposed by college administra tor r eopeni ng decisions, not just on coll ege-associat ed populati ons, but on vulne rabl e individual s in surrounding communities. Al though ou r primary focus has been on using viral gene tics to tr ac k introductio n and spread of th ese substr ains, we not e th at the advers e effects of college-amplified c lusters ar e not limi ted to the mo rbidity and mor tali ty of infecte d individuals and the subs tanti al associat ed heal thcare cos ts. Rapid case growth, as exp erienc ed in La Crosse, intensifies the magnitude of the mu ltifacete d adverse effects of the pand emic upon communiti es. These effects includ e significantly ext ending the p eriod during which children are un able to at te nd in-person K-12 schooling (19), further exace rba tion of the many psychosocial stresses affecting bot h individuals and families (20), extending the ongoing unemployment crisis (21), placing more families at risk of losing healt h insurance ( 22) and harming small . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 9 businesses (23). These factors should all be consider ed as par t of any cost-benefit analysis of reopening colleges and universi ties.

This work was supported by the G unders en Medical Fou ndati on. PK holds the Dr. Jon & Bet ty Kabara Endowed Chair in Precision Oncology. W e thank th e members of th e Gund ersen Medical Found ation ’s Molecular Diagnos tics Laborat ory for pro viding the specimens used in this st udy.

1. Zhu N, Zhang D, Wang W, Li X, Yang B, Song J , et al. A Novel Coron avirus from Pati ents with Pneumonia in China, 2019. N Engl J Med 2020 ;382:727-33 2. van Dorp L, Acman M, Richard D, Shaw LP, Ford CE, Ormond L , et al. Eme rgence o f genomic diversity and recurren t mu ta tions in SAR S-CoV-2. Infect Gene t Evol 2020 ;83:104351 3. US News and Wor ld Repo rt . . 4. Li H, Handsaker B, Wysoke r A, Fenn ell T, Ruan J, Hom er N , et al. Th e Seque nce Ali gnment/Map format and SAM tools . Bioinforma tics 2009 ;25:2078-9 5. Kim D, Pag gi JM, Park C, Bennet t C, Salzb erg SL. Grap h-based genom e alignment a nd genotyping with HISAT2 and HIS AT-genotype . Na t Bi otechnol 2019 ;37 :907-15 6. Cavener DR. Comparison of the cons ens us sequence flanking tra nslatio nal sta rt s ites in Drosophila and ver tebr at es. Nucl eic Acids Res 1987 ;15:1353-61 7. Shu Y, McCauley J. G ISAID: Global ini tiati ve on sharing all influenza da ta - from vision to re ality. Euro Surveill 2017 ;22 8. Hadfield J, M egill C, Bell SM, Huddl eston J, Pott er B, Callen der C , et al. N ex tstr ain: real-time tracking of pathogen evolution . Bioinfor matics 2018 ;34:4121-3 9. Richmond CS, Sabin AP, Jobe DA, Lovrich SD, Kenny PA. Inter regional S ARS-CoV-2 spread from a single introduc tion ou tbre ak in a meat-p acking plant in north eas t Iowa. med Rxiv 2020 :2020.06.08.20125534 10. Chaudhuri G, Oxley S, W enzlaff S. Mappi ng Spatiot emporal Pa tte rns of Liquor Law Violation Citations During Okto berfes t in College Town of La Crosse, Wisconsin. Forensic GI S: Springer ; 2014. p 201-20. 11. Swanson DJ, Zegers KM, Zwaska AA. Impl ementing a social no rms approach to re d uce alcohol abuse on campus: Lessons le arned in the shadow of ‘The World's Larg est Six-Pack’ . The Social Science Journal 2004 ;41:621-35 12. Boehmer TK, DeVies J, Carus o E, van Sant en KL, Tang S, Black CL , et al. Changing Age Distribution of the COVID-19 Pandemic - United Stat e s, May-August 2020. MMWR Morb Mo rt al Wkly Rep 2020 ;69:1404-9 13. Salvator e PP, Sula E, Coyle JP, Caruso E, Smith AR, Levine RS , et al. R ecen t Incr eas e in COVID-19 Cases Repor ted Among Adul ts Aged 18-2 2 Years - United St at es, May 31-Sept ember 5, 2020. MMWR Mo rb Mor tal Wkly Rep 2020 ;69: 1419-24 . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 10 14. Paltiel AD, Zheng A, Wal ensky RP. Assessment of SARS-CoV-2 Screening Str ategi es to Permit th e Safe Reop ening of College Campuses in the Unit ed Sta tes. JAM A Ne tw Open 2020 ;3:e2016818 15. Balli J. “I felt like I was in a lot of danger”: UWL students tell all on qu aran tine in W entz Hall . The Raque t Press 2020; htt ps://ther acquet .or g/10608/news /i-felt-like-i-was-in -a-lot-of-danger-uwl- studen ts-tell-all-on-qua ranti ne-in-wentz- hall/ . 16. Wilson E, Donovan CV, Campbell M, Chai T, Pittman K, Sena AC , et al. Mul tiple CO VID-19 Clusters on a Universi ty Campus - North Carolina, Augus t 2020. MMWR Morb Mo rtal Wkly Rep 2020 ;69:1416-8 17. Yamey G, Walensky RP. Covid-19: re-ope ning universities is high risk. BM J 2020 ;3 70:m3365 18. Willoughby T, Good M, Adachi PJ, Hamza C, Tavernier R. Examining the link be twe en adolesce nt brain developm ent an d risk taking from a social-developmen tal pe rspective (rep r inted). Br ain Cogn 2014 ;89:70-8 19. Masonbrink AR , Hurley E. Advoca ting for Children During the COVID-19 School Closures. Pediatrics 2020 ;146 20. Pfefferbaum B, Nort h CS. Ment al Heal th and the Covid-19 Pandemic. N Engl J Me d 2020 ;383:510-2 21. Blustein DL, Duffy R, Ferreira J A, Cohen-S cali V, Cinamon RG, Allan B A. Un employment in th e time of COVID-19: A research ag enda . J Vocat Beh av 2020 ;119:103436 22. Woolhandl er S, Himmels tein DU. In te rse cting U.S. Epidemics: COVID-19 and Lack of Health Insurance . Ann I nte rn Med 2020 ;173 :63-4 23. Bartik A W, Be rtr and M, Cullen Z, Gl aese r EL, Luca M, Stanton C. The impac t of COVID-19 on small business outcomes and expec ta tio ns. Proc Natl Aca d Sci U S A 2020 ;117:17 656-66 . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint 11 FIGURES LEGENDS Figure 1. Cumulative SARS-CoV-2 case in cidence in La Crosse County, WI , stra tifie d by age group (March – Sept ember 2020). Figure 2. Choropleth hea tmaps showing a weekly assessment of the 7-day rolling average of daily case numbers per 100,000 pop ulati on in cens us tracts in th e city of La Crosse. Census t ract popul ations wer e taken from 2010 census data . The loca tio ns of University of Wisconsin – La Crosse (UWL), Western Technical College (WTC) and Viterbo Uni versity (VIT) are indicated. Figure 3. Radial time resolved phyloge ne tic tre es containi ng (A) all 514 high-quality genomes sequenced betwee n 3/18/20 and 9/28/20 and (B) hi ghlighting just those specimens seq uenc ed in Sept ember . Sequencing resul ts ar e from a 21-county region of Wisconsin, Minn esot a and Iow a. The case growth in La Crosse County in Septemb er was att ri buted t o the Coll ege A and College B clus ters. Figure 4. Phylogenetic trees demons tra ti ng the rela tionships b etwe en cases in th e College A and College B substrains . Trees a re scaled by sequ ence divergence so th at id entical gen omes a re vertically align ed and each furth er incr ement to th e right r epres ents acquisi tion a new nucleo tide v ariant . Clade-defining mutations a re indica ted in bo th cases. Gr oups of 5 or more gene tically identica l cases are highlight ed in orange, with the medi an age of individua ls in each group indicat ed. Transmission chains in two skilled nursing facilities, de tect ed in th e College B cluster, a re also highligh ted. . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint . CC-BY-NC-ND 4.0 International licenseIt is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted October 14, 2020. ; https://doi.org/10.1101/2020.10.12.20210294doi: medRxiv preprint