Since the first patient was diagnosed with COVID-19 in Lombardy, on February 20 th 2020, Italy has become the third most affected country (>215.000 cases) and 30 .000 deaths in the world, as of May 8 th , 2020. [1] COVID-19 neurotropic properties may underlie a worsening of chronic neurological diseases, such as Parkinson's disease (PD). COVID-19 may worsen PD by a number of mechanisms, [2] [3] [4] including pharmacodynamics changes (e.g., reciprocal interactions between the dopaminergic and reninangiotensin systems in the substantia nigra and striatum [5] ) as well as systemic inflammatory response. [6] [7] [8] [9] Patients with PD are frailer than general population because of disease-related factors and age-related co-morbidities. [2-4;10,11] A higher COVID-19 mortality rate has been described in advanced PD patients in association to older age and longer disease duration. [12] Our primary objective was to investigate the effects of COVID-19 on motor and nonmotor symptoms in a community-based PD cohort. In addition, we explored whether older age and longer disease duration represented risk factors for developing symptomatic COVID-19.
In the present observational, community-based, case-control study, we investigated demographic and clinical features in a cohort of patients with idiopathic PD [13] and COVID-19 as compared to control PD subjects between the pre-outbreak period in Italy (January 01 st , 2020) and the end of lockdown restrictions (May 4 th , 2020). A 3-month period was chosen to minimize the effect of PD progression on the change in clinical features and the recall bias. Diagnosis of COVID-19 was performed according to clinical and laboratory criteria for probable and confirmed cases, released by the World Health Organization criteria on March 20 th , 2020. [14] In symptomatic cases fulfilling criteria for 'probable case' [14, 15] we included only patients with close and protracted contacts (i.e., caregivers) with laboratory-confirmed cases.
Out of the 1092 records obtained by searching the Besta Institute clinical software for patients fulfilling the following criteria (i) International Classification of Diseases, Ninth-Revision, Clinical Modification code for parkinsonism 332.0, (ii) resident in the Lombardy region, northern Italy (which is by far the most affected area (>80.000 cases) with the highest case-fatality (>15.000 deaths) in Italy, as of to May 11 th , 2020), [16] (ii) visited at least once from January 01 st , 2019 to December 31 st , 2019, we performed a random selection of 150 PD for subsequent remote interview by a neurologist experienced in movement disorders (by video-consultation or telephone), [2, 17, 18] which was performed between April 15 th and May 4 th , 2020. Signed informed consent was obtained prior to remote assessment and collection of clinical data, as approved by the local Ethics Committee.
Out of a total of 141 subjects who accepted to be interviewed, twelve (8.5%) were affected by COVID-19. [14] Compared to overall cohort of 129 PD control subjects (males 56.6%; age 69.1±10.1 years; PD duration 8.2±5.0 years; any comorbidity 62.8%), cases has similar sex distribution (P=0.37), age (P=0.24), PD duration (P=0. 20) , and co-morbidities (P=0.54). Among those 129 screening negative for COVID-19, a group of 36 PD controls matched for sex, age and disease duration (±1 year) was used for subsequent statistical analysis. A 1:3 ratio was chosen to minimise the effects related to biological variability and the potential presence of asymptomatic COVID-19 among controls. [19] Cases and matched controls underwent in-depth assessment using the following internationally-validated scales.
Motor aspects of experiences of daily living and the severity of treatment-related motor complications were assessed using the Movement Disorders Society Unified PD Rating Scale (MDS-UPDRS) parts II and IV, respectively; [20] non-motor symptoms (NMS) were assessed using of the Italian version of the NMS Scale (NMSS); [21] overall changes of motor and nonmotor features was additionally rated using the Clinical Impression of Severity Index for Parkinson's Disease (CISI-PD). [22] Descriptive statistics were provided for continuous (mean and standard deviation) and categorical (count and percentage) variables, which were compared between groups using the Student's t-test and the Fisher's exact test. Then, between-visit changes in study parameters (January-to-April, 2020) were compared between COVID-19 cases and controls using repeated-measure linear regression models. The role of potential confounders was addressed by MANOVA. All statistical analyses were performed using STATA statistical software release 15.1 (Stata Corporation, College Station, TX) setting the level of significance at a two-tailed P-value <0.05.
Demographic and clinical features at baseline were similar between PD cases and matched control subjects, except for the greater need to increase dopaminergic therapy dosing and higher rate of contacts with confirmed COVID-19 among cases ( Table 1) . COVID-19 symptoms were mild, managed at home without symptomatic therapy in 3 cases (25%); 8 cases (66.7%) had moderate illness, pharmacologically managed at home by the general practitioner; only one patient was hospitalized (8.3%) due to pneumonia. COVID symptoms remitted in ten out of 12 (83.3%) cases and were still ongoing in two patients (16.7%; both remitted at subsequent follow-up on May 15 th ); nobody died.
At within-group comparison, cases evidenced a significant worsening of the CISI-PD total and motor-signs scores, MDS-UPDRS part-II score, the NMSS total score and the urinary domain sub-score ( Table 2 , footnote d). Between-group case-control analysis additionally revealed greater motor disability (at CISI-PD), motor fluctuations (at MDS-UPDRS part-IV), and nonmotor complaints.
Involvement of cognitive functions was marginal (no change at CISI-PD), and autonomic cardiovascular and sexual functions remained unaffected ( Table 2) .
Finally, considering the greater rate of diarrhoea among COVID-19 cases ( Table 1 ) and its detrimental effect on the pharmacokinetics of dopaminergic medications (particularly levodopa), we adjusted a selected subset of models for this symptom and found that worsening of CISI-PD total and motor signs scores was partially mediated by diarrhoea (p=0.002 for both) although COVID-19 status was still a significant contributor (p=0.025 and 0.026, respectively). Worsening of MDS-UPDRS part II and the part IV total score and the NMSS total score were explained by COVID-19 alone (p=0.008, p=0.034, p=0.008, respectively); interestingly, increase in daily OFF-time was fully explained by diarrhoea (p=0.019). We additionally explored whether urinary dysfunction and fatigue were due to COVID-19 or to diminished dopaminergic drive ( Table 2 ) and found that urinary problems worsening was due by both motor fluctuations (p<0.001) and COVID-19 (p=0.005), while fatigue was due to COVID-19 alone (p<0.001).
To our knowledge, this is the first community-based case-control study describing the effects of symptomatic COVID-19 on PD motor and nonmotor symptoms. First, PD patients who develop symptomatic COVID-19 were neither older age nor with longer disease duration than those screening negative, but rather had nonsignificant 3.5-year younger age and 2.0-year shorter disease duration. The lack of case fatalities is consistent with the Italian case-fatality rate of 3.5% at 60-69 years of age. [15] Similarly, only one patient (8.3%) needed to be admitted to hospital for severe COVID-19 illness, which is in line with the frequency of hospitalization previously reported in general population of the Lombardy region. [23] We expand previous report in advanced PD [12] and show that mild-to-moderate COVID-19 may be contracted independently of age and PD duration and that PD patients with midstage PD do not seem to have an overall worse outcome than non-PD population. [15] Concerning the primary objective of the study, we found a worsening of motor and nonmotor symptoms of PD in the COVID-19 group compared to matched control subjects over the study period.
Motor symptoms. COVID-19 induced a significant worsening of motor performance, motorrelated disability and experiences of daily living. Worsening of levodopa-responsive motor symptoms and increased daily OFF-time, caused either by the effects of acute systemic inflammatory response [6] [7] [8] [9] or by changes in pharmacokinetics, was so pronounced in one-third of cases to prompt neurologists to increase dopaminergic therapy. We explored the relative contribution of suboptimal absorption of oral therapy by adjusting motor endpoints for diarrhoea, which was present in 50% of cases. According to multivariate analysis, concomitant diarrhoea explained the increase in motor fluctuations, but could not entirely explain the worsening of motor disability and motor aspects of experiences of daily living.
Nonmotor symptoms. COVID-19 significant aggravated a number of nonmotor symptoms. Increased fatigue in our cohort was fully explained by COVID-19, confirming that it a common COVID-19
symptom [24] , as described in PD following systemic inflammation. [25] Urinary urge/incontinence and nicturia were explained by the infection as well as increased motor fluctuations, which were partly due to pharmacokinetic issues. COVID-19 was neither a major cause of cognitive dysfunction nor autonomic failure in our cohort of mid-stage PD. Although there was an effect of COVID-19 on attention, it was not severe enough to be detected by CISI-PD. We neither found changes in cardiovascular, gastrointestinal and sexual function domains at the NMSS nor differences in hypotension between the groups.
The main limitation of this study is the small cohort of COVID-19 patients, albeit statistical analysis could detect several significant changes. There are a number of strengths worth mentioning. First, our case-control study protocol excluded the detrimental effects that quarantine and lockdown restrictions might have played on a number of confounders that influence PD motor and nonmotor symptoms, such as reduced physical activity, enhanced stress, confusion, anxiety, and sleep disturbances. [26] Second, a community-based survey minimised selection bias related to the inclusion of hospitalized patients with severe illness or institutionalised ones with advanced PD and comorbidities, who are more susceptible to neurological manifestations worse outcome. [27, 28] Our cohort with mild-to-moderate illness is likely representative of the majority of PD affected by COVID-19, considering that only a minority of cases require hospitalization [29] [30] [31] . Third, we excluded secondary and atypical parkinsonisms (including dementia with Lewy bodies), minimising the risk of overestimating either fluctuating or drug-induced worsening of cognitive dysfunction, visual hallucinations, and autonomic failure. Finally, our comprehensive assessment performed by experienced neurologists in a single tertiary referral clinic ensured a homogeneous and standardized approach.
Larger multicentre studies would have requested longer time for data collection, increasing patients/caregivers recall bias.
PD patients may experience substantial worsening of motor and nonmotor symptoms during mild-tomoderate COVID-19 illness, independently of age and disease duration. Clinicians should take pharmacokinetic changes into consideration before adjusting therapy regimen (e.g., management of dehydration secondary to fever, diarrhoea, anorexia with reduced water intake). Although we speculate that subacute clinical changes in PD associated to nonsevere COVID-19 illness are likely caused by systemic inflammatory response [8] [9] rather than a direct invasion of the central nervous system, [27, 28] further studies in larger PD populations are warranted to clarify the cause-effect relationship among clinical changes and the severity of COVID-19 illness, cytokine levels and virus detection in the cerebrospinal fluid. 