on Infectivity of Air on a Hospital Ward

Transcription

1 Materials for Online Data Supplement for AJRCCM Surgical Face Masks Worn by Patients with Multidrug-Resistant Tuberculosis: Impact on Infectivity of Air on a Hospital Ward Authors: Ashwin S. Dharmadhikari 1,2, Matsie Mphahlele 3, Anton Stoltz 4, Kobus Venter 3, Rirhandzu Mathebula 3, Thabiso Masotla 3, Willem Lubbe 3, Marcello Pagano 5, Melvin First 6, Paul A. Jensen 7, Martie van der Walt 3, Edward A. Nardell 2,1 Detailed Methods AIR Facility The Airborne Infections Research (AIR) facility was designed to expose susceptible sentinel guinea pigs to airborne tuberculosis (infectious droplet nuclei) generated by patients in order to test transmission control interventions. Patients in the ward are those with active, culture-proven, mostly sputum smear positive, pulmonary MDR tuberculosis receiving standardized MDR-TB treatment. Guinea pigs are an ideal model to study TB transmission given their exquisite susceptibility to infection and delayed type hypersensitivity reaction to purified protein derivative (PPD) after airborne exposure and infection. The AIR facility is part of the Mpumalanga provincial MDR-TB referral hospital and consists of a 6-bed inpatient MDR-TB ward connected to 2 guinea pig exposure chambers by an airtight ventilation system. HEPA filter treated air enters and infectious air leaves the patient ward at a rate of 6 room volume air changes per hour (ACH) and is then delivered entirely to one or the other of two guinea pig exposure chambers (Figure 1 in main manuscript).

2 Within each guinea pig exposure chamber, the air being delivered from the patient ward emerges from a grid of 60 individual ducts located on a side wall. Each of these 60 ducts supplies ward air to one cage of animals (1:1 duct to cage ratio). Each cage held two animals, and for this study, only 45 of the 60 cages in each animal chamber were occupied by guinea pigs (total 90 guinea pigs in each side). The cages are located 1 cm from the opening face of the duct supplying that cage. Air from the guinea pig chamber is then actively exhausted through a single exhaust duct located on the ceiling. The direction of air flow ensures that air emerges from the inlet duct and passes through the guinea pig cages prior to exiting the animal exposure chamber. In addition, an electronic building monitoring system measured the airflow velocity at specific locations in the supply and exhaust duct work of the patient wards and each animal chamber, as well as the humidity and temperature of the air several times each day during the study. These measurements ensured that airflow, temperature, and humidity were the same to each group of guinea pigs on mask-on and mask-off days. In addition, there was no infiltration of any outside air into the guinea pig exposure chambers. The guinea pig exposure chambers were separated from the corridor by an anteroom. The anteroom was at negative pressure relative to the corridor and at neutral pressure relative to the guinea pig exposure chambers. In this study, guinea pigs in each chamber sampled ward air between 7 AM and 7 PM under one of two experimental conditions. On non-mask days the ward air was exhausted to the control guinea pig chamber from 7 AM to 7 AM the following day, but

3 between 7 PM and 7 AM, ward air was irradiated with high intensity in-duct ultraviolet germicidal irradiation to eliminate infectious particles prior to passage through the guinea pig exposure chamber and facilitate guinea pig exposure to aerosols only during the 12 hour period from 7 AM 7 PM. On mask days, the ward air was exhausted to the intervention guinea pig chamber from 7 AM to 7 AM the following day and irradiated between 7 PM and 7 AM as described. On days when animals in a given chamber were not exposed to ward air, they received a separate supply of HEPA filtered, conditioned air that bypassed the patient ward. Patient enrollment and selection Seventeen patients (8 males, 9 females) occupied the AIR Facility ward during the study. These patients were sequentially selected from among patients newly admitted for hospitalization and MDR-TB treatment initiation at the adjacent hospital between May August Patients had to have culture proven MDR-TB and cough. Most (82%) were also sputum smear positive for acid fast bacilli. They were approached for enrollment only if they were ambulatory, could tolerate mask wearing for extended periods of time, and had no contraindications to mask usage (e.g. room air oxygen saturation < 92%, requirement for supplemental oxygen, severe obstructive lung disease, or respiratory insufficiency that might be worsened by mask usage). Informed consent was obtained from each patient prior to enrollment. Patients contributed equal amounts of time under control and intervention conditions to each guinea pig cohort. All patients received MDR-TB treatment in accordance with South African guidelines (1).

4 Clinical and laboratory characteristics of the patient cohort are summarized in Table 1 in the main manuscript. Study protocol Upon admission, patients were given instructions on how to wear the surgical mask. Mask usage occurred on alternate days during the entire study period of 3 months (12 weeks). On mask use days, patients were instructed to wear the surgical mask over their mouth and nose from 7 AM 7 PM, for at least 80% of the time. Masks could be removed for meals and medications. Masks were not worn during sleep or between the hours of 7 PM 7 AM. As described earlier, the in-duct ultraviolet irradiation of ward air between 7 PM 7 AM meant that guinea pigs sampled infectious ward air between 7 AM 7 PM. Mask use was monitored by nursing staff through 10 daily unannounced spot checks on each of their patients. The spot checks occurred approximately every 2 hours. Patients in the study were informed at study entry that they would be monitored for mask usage. Those who demonstrated the highest levels of adherence to the study protocol were eligible for a small non-monetary incentive (e.g. additional snacks/beverages) at the conclusion of the study. Guinea Pigs and Tuberculin Skin Testing Outbred male and female, specific pathogen free, Dunkin-Hartley guinea pigs (National Health Laboratories Services, South Africa; n = 180; 90 per exposure chamber) were acquired at 6 weeks of age weighing g. All animal husbandry and experimental procedures were executed in accordance with institutionally approved

5 protocols. Tuberculin skin testing (TST) was performed with 100 tuberculin units (2.2 µg) per 0.1 ml PPD (Orme Lab, Colorado, USA). PPD was diluted immediately before testing in phosphate buffered saline containing 0.05% Tween 80 (v/v) and was administered each time on a different depilated area of the back via intradermal route. Animals underwent monthly TSTs before, during and 1 month after MDR-TB exposure ended, with the final test used to detect infections that may have occurred in the last few weeks of the exposure period (2). TSTs were read in a blinded, duplicate manner using digital calipers (Wilson Wolpert, The Netherlands). Two independent readings were taken at right angles to each other (longitudinal and transverse) at 24 hours postadministration by a single animal handler and the average value was used for analysis. Any induration was considered indicative of infection. We also examined whether a threshold value of 6 mm induration (to potentially improve test specificity and based on prior work with guinea pigs naturally exposed to TB aerosols (3)) would make a difference in our results. When we used a higher TST induration threshold value of 6 mm for our calculations, we found similar results regarding efficacy (results not shown). The change in the threshold from 0 mm to 6 mm only resulted in the reclassification of 3 guinea pigs in the intervention group as not infected. However, this change did not significantly alter the difference in the risk of infection between the groups (results not shown for analyses using 6 mm threshold). In addition, these pathogen free guinea pigs were not exposed to environmental mycobacteria, making any reaction to PPD specific to M. tuberculosis infection acquired during the study.

6 Animals exhibiting a reactive TST were removed from the exposure chambers and housed in a separate animal facility for observation until they were euthanized. In addition, a completely separate group of uninfected guinea pigs (n=6) and guinea pigs infected intranasally with M. tuberculosis H37Rv (n=6) served as positive and negative TST controls for PPD reagent. They underwent TSTs at the same interval as ward air exposed animals. References Cited in Online Supplement E1. South africa department of health: Management of drug-resistant tuberculosis - draft policy guidelines E2. Smith DW, McMurray DN, Wiegeshaus EH, Grover AA, Harding GE. Hostparasite relationships in experimental airborne tuberculosis. Iv. Early events in the course of infection in vaccinated and nonvaccinated guinea pigs. Am Rev Respir Dis 1970;102: E3. Dharmadhikari AS, Basaraba RJ, Van Der Walt ML, Weyer K, Mphahlele M, Venter K, Jensen PA, First MW, Parsons S, McMurray DN, Orme IM, Nardell EA. Natural infection of guinea pigs exposed to patients with highly drug-resistant tuberculosis. Tuberculosis (Edinb) 2011.