Surface Cleaning and Disinfection: Efficacy Assessment of Four

Subscriber access provided by University of Newcastle, Australia

Article

Surface cleaning and disinfection: efficacy assessment of four chlorine types using E. coli and the Ebola surrogate Phi6 Karin Gallandat, Marlene K. Wolfe, and Daniele S. Lantagne Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.6b06014 • Publication Date (Web): 15 Mar 2017 Downloaded from http://pubs.acs.org on March 16, 2017

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Environmental Science & Technology is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 29

Environmental Science & Technology

1

Surface cleaning and disinfection:

2

Efficacy assessment of four chlorine types

3

using E. coli and the Ebola surrogate Phi6

4

Karin Gallandat1*, Marlene Wolfe1, and Daniele Lantagne1

5

1: Department of Civil and Environmental Engineering, Tufts University, Medford, MA, USA

6

*Corresponding author: 200 College Ave, Medford, MA 02155, USA.

7

E-mail: [email protected]. Phone: +1 617 483 2045.

8

ACS Paragon Plus Environment


9

ABSTRACT

10

In the 2014 West African Ebola outbreak, international organizations provided conflicting

11

recommendations for disinfecting surfaces contaminated by uncontrolled patient spills. We

12

compared the efficacy of four chlorine solutions (sodium hypochlorite, sodium

13

dichloroisocyanurate, high-test hypochlorite, and generated hypochlorite) for disinfection of

14

three surface types (stainless steel, heavy duty tarp, and nitrile), with and without pre-cleaning

15

practices (pre-wiping and/or covering) and soil load. The test organisms were E. coli and the

16

Ebola surrogate Phi6. All tests achieved a minimum of 5.9- and 3.1-log removal in E. coli and

17

Phi6, respectively. A 15-minute exposure to 0.5% chlorine was sufficient to ensure 0.05), except on heavy-duty tarp with soil load (p = 0.03).

appeared

equally

efficacious,

there

was

no

significant

difference

between

274

A post-hoc analysis was conducted using Dunn’s test to compare chlorine types and

275

recommendations respectively for the two cases where statistically significant differences were

276

detected but no chlorine type or recommendation appeared to systematically perform better than

277

the others (results not shown).



Page 14 of 29

278

Only NaDCC was used to evaluate the effect of a 15-minute exposure time and of a towel

279

soaked in chlorine solution, as significant differences between chlorine type were not routinely

280

identified. On heavy-duty tarp, increasing exposure time from 10 to 15 minutes did not

281

significantly increase efficacy in any of the four recommendations with and without soil load, the

282

only exception being in presence of soil load when doing nothing before applying chlorine

283

(WMW test, p = 0.03). Applying a towel soaked in 0.5% NaDCC solution on the spill was not

284

significantly more efficacious than using a dry towel to cover the spill on heavy-duty tarp

285

(WMW test, p >0.05 both with and without soil load).

286 287

Phi6 results.

288

The surface carriers were inoculated with 8·107 CFU on average (range 1.3·106 to 2.3·108

289

CFU). The recovery rates, as estimated based on the positive controls after 1-hour drying, were

290

on the order of 20% on all three surfaces.

291

All recommendations achieved at least 3-log Phi6 removal (Table 3). Out of a total of 288

292

samples, Phi6 was detected after disinfection in six samples on nitrile and in no samples on

293

stainless steel or heavy-duty tarp. The residual contamination in the positive samples ranged

294

from 16 PFU/cm2 to 80 PFU/cm2 after disinfection. Due to the limited number of positive

295

samples, statistical tests were therefore not conducted on Phi6 results.

296

The geometric mean of triplicates exceeded the target of 10 residual PFU/cm2 in two instances

297

(Figure 4): Phi6 was detected in presence of soil load when pre-cleaning nitrile discs with both

298

generated and stabilized NaOCl.

299 300

For the two conditions where Phi6 was detected on nitrile after a 10-minute exposure to 0.5% chlorine, increasing the exposure time to 15 minutes led to all non-detectable levels.


Page 15 of 29


301

DISCUSSION

302

We conducted a systematic evaluation of the efficacy of surface disinfection recommendations

303

for Ebola contexts, including testing two organisms, two soil load conditions, four surface

304

cleaning recommendations, and four chlorine types on three surfaces commonly used in ETUs.

305

Results indicated that test organism, surface type, and covering spills influenced surface

306

disinfection efficacy, while chlorine type, soil load, and pre-cleaning did not. Overall, the

307

recommendations provided during the 2014 EVD outbreak for surface disinfection appeared to

308

be efficacious at removing E. coli and Phi6 from representative surfaces. Based on our results,

309

we recommend a 15-minute exposure time to 0.5% chlorine to ensure safe surface disinfection in

310

EVD contexts. These results: 1) are consistent with previous results while also providing further

311

context-specific evidence; 2) answer questions about chlorine type, while raising questions about

312

the role of soil load and how to appropriately model soil load in these contexts; 3) highlight the

313

important role of test organism and surface type in disinfection; and, 4) inform how to conduct

314

research on surface disinfection efficacy in further outbreak situations.

315

The Cook et al.23,25 studies documented a 5-minute contact time with 0.5% sodium

316

hypochlorite to completely disinfect stainless steel surfaces inoculated with the Ebola virus. Our

317

data expand on their work as we found that 10 minutes were sufficient to disinfect stainless steel

318

using Phi6 as an Ebola surrogate, but additional contact time was needed specifically on nitrile to

319

ensure full disinfection under all tested conditions. This is also consistent with a previous study

320

where environmental sampling was conducted in an isolation ward in Uganda in 2000. Bausch et

321

al.13 found that CDC and WHO recommendations were efficacious, as Ebola was not isolated

322

from surfaces after routine disinfection. However, the recommendations were more conservative

323

during that time, as they recommended a 1% chlorine solution with a 15-minute contact time,



Page 16 of 29

324

and that a 10% chlorine solution should be used for heavy or dense spills.37 A more recent study

325

conducted in Sierra Leone in 2015 suggests that environmental decontamination using 0.5%

326

sodium hypochlorite was mostly effective at removing Ebola RNA from surfaces around the

327

bedside, where contamination was detected most often.11 Our results also indicate 0.5% chlorine

328

solutions should be efficacious.

329

In contrast to beliefs that some chlorine types may be more efficacious than others, we found

330

all chlorine types appeared equally efficacious for the disinfection of surfaces against E. coli and

331

Phi6. We attribute this to the high chlorine dose (0.5%) applied. As such, we recommend using

332

whichever chlorine source is available, safe to handle, and maintains an appropriate

333

concentration.38

334

The presence of soil load did not affect disinfection efficacy at a chlorine concentration of

335

0.5%. We used the soil load mixture recommended in the ASTM International Standard26 and the

336

chlorine demand of the spill with soil load represented less than 0.01% of the amount of applied

337

chlorine. Again, we hypothesize soil load did not impact efficacy because of the high chlorine

338

concentration applied. While this is likely representative of field practices, where chlorine will

339

be generously applied to ensure disinfection, further research is needed to develop new matrices

340

more representative of liquid Ebola bodily wastes and their impact on disinfection efficacy.

341

Our results suggested that test organism and surface type do play an important role in

342

determining disinfection efficacy. We found that E. coli was more challenging to disinfect on

343

heavy-duty tarp, and Phi6 was more challenging to disinfect on nitrile; suggesting that different

344

mechanisms are responsible for disinfection of the two organisms. A literature search provided

345

little information on which fundamental mechanisms might explain the observed difference. We

346

hypothesize that physical properties of the surface (such as roughness) as well as surface charge


Page 17 of 29


347

(of both the surface and the test organism) impacted disinfection efficacy. The fact that

348

increasing the exposure time from 10 to 15 minutes did not improve disinfection efficacy against

349

E. coli on heavy duty tarp suggests that bacteria did not enter in contact with the disinfectant,

350

possibly due to some physical protection provided by the roughness of heavy duty tarp. It is

351

unclear why such an effect would not be observed with Phi6, and additional research is indicated

352

to investigate our hypotheses. The widespread use of both heavy-duty tarp and nitrile in

353

emergency settings makes these results of particular concern. It is recommended to conduct

354

future research with other surfaces commonly found in ETU settings, including paper, cardboard,

355

cotton, rubber, latex, and concrete.

356

The international recommendations for surface disinfection in Ebola contexts are particularly

357

different in terms of practices. It has been suggested to wipe and to cover spills, and recently, to

358

pre-clean surfaces with water and detergent.39 While pre-cleaning reduces the volume of spill to

359

be disinfected, such a practice generates contaminated waste and can result in exposure of

360

healthcare personnel to infectious materials. Although we were not able to test for the

361

multiplicity of pre-cleaning practices that are likely to be encountered in the field (with or

362

without soap, with dry and wet towels, and accounting for individual variability), our results

363

suggest that wiping the surface before applying chlorine is unlikely to increase disinfection

364

efficacy. This is consistent with a study of surface disinfection involving several viruses, where

365

Tuladhar et al,.12 found that inactivation by a 0.1% chlorine solution was proportionally more

366

important than the action of wiping. Covering spills is recommended to avoid disease

367

transmission by splashes18 and our results with Phi6 suggest that it does not affect disinfection

368

efficacy. We recommend covering spills with a cloth soaked in 0.5% chlorine and leaving it for



369

15 minutes if disease transmission via splashes is a concern, as is the case with EVD due to the

370

very low infectious dose.

Page 18 of 29

371

Our study had limitations, including that: 1) the comparison of E. coli and Phi6 is limited

372

by different starting concentrations and detection limits; 2) although the recovery rates were

373

similar for both organisms, we cannot rule out that some of the observed differences could be

374

due to variability in recovery; 3) we used a surrogate organism instead of the actual virus, albeit

375

one selected based on a comparison between the surrogate (tested in our laboratory) and Ebola

376

virus (tested by Cook et al.23 in a Biosafety Level 4 laboratory); 4) we believe that our protocol,

377

using 8-cm in diameter discs instead of the 1-cm diameter surface carriers recommended in the

378

ASTM Standard,33 simulates field conditions more accurately, however 2-mL spills are still

379

unlikely to reflect the extent of environmental contamination experienced in ETUs; 5) our

380

standardization of pre-cleaning recommendations does not account for further variability in pre-

381

cleaning practices applied in ETU settings; 6) the ASTM soil load might not be representative of

382

the matrices in which the Ebola virus would be shed by patients; and, 7) temperature and relative

383

humidity in the laboratory were both lower and more controlled than those commonly found in

384

ETU settings.

385

Despite these limitations, we feel our data contribute to the understanding of, and assist in

386

providing an evidence base for, surface disinfection practices in outbreaks. In particular, our

387

results support the recommendation of a 15-minute exposure to 0.5% chlorine – independently of

388

chlorine type, surface type, practices, and presence of organic matter – as an efficacious measure

389

to interrupt EVD transmission via fomites. Using Phi6 as a surrogate allowed us to carry out

390

extensive testing and to identify critical conditions; we recommend evaluating the resistance of

391

the Ebola virus (at Biosafety Level 4) to a 15-minute exposure to 0.5% chlorine, without pre-


Page 19 of 29


392

cleaning or covering, with or without soil load, on nitrile and, if possible, on other surfaces, to

393

confirm the results presented here.

394

Further research is required to investigate how surface properties and complex matrices affect

395

disinfection efficacy, as well as to understand the role of wiping or covering spills. A more

396

fundamental understanding of the mechanisms affecting disinfection efficacy and of the physico-

397

chemical interactions between microorganisms and surfaces will allow extension of these results

398

to different pathogens with more flexibility.

399

Our study was carried out in response to the 2014 Ebola outbreak and focused on evaluating

400

the efficacy of existing recommendations in place during that outbreak. It is recommended future

401

research be completed to develop appropriate recommendations, not just test the efficacy of

402

existing recommendations. For instance, using NaDCC granules instead of a liquid chlorine

403

solution to cover uncontrolled spills has been identified as an efficacious procedure against

404

bacteria in developed country hospitals.29 Additional research is needed to evaluate the adequacy

405

of this, and other novel, protocols as a possible intervention in ETU contexts.

406

For diseases with infectious doses as low as EVD, multi-barrier approaches (including

407

personal protective equipment, handwashing, disinfection of surfaces and safe waste

408

management) should be used to minimize transmission risk. However, multi-barrier approaches

409

– particularly those with waiting times – are burdensome in the ETU context, due to the

410

maximum time responders can spend in PPE. A broader reflection around the best way to

411

minimize transmission risk and the appropriate number and level of barriers to use in healthcare

412

facilities and communities facing situations such as EVD is necessary. For example, it has been

413

suggested to depend on PPE primarily, and not be concerned with disinfecting spills in the ETU

414

context. Ideally, further discussion will involve responders, members of the scientific community



415

and international organizations to address the appropriateness of existing recommendations and

416

how they can be adjusted to increase preparedness for future outbreaks.


Page 20 of 29

Page 21 of 29


417

TABLES

418

Table 1: Recommendations on surfaces disinfection in Ebola outbreaks.

419 420

Target

Action

Disinfectant

Exposure time

Source

Hospital/ETU

Pre-clean surface

0.5% chlorine

10 minutes

WHO19

Household

Cover spills

0.5% chlorine

15 minutes

CDC40

Hospital

Pre-clean surface

“Chemical disinfectant for non-enveloped viruses”

Not specified

CDC20

ETU

Do nothing

0.5% chlorine

15 minutes

MSF18

Table 2: Chlorine types commonly used in emergency contexts.

Chlorine type

Expected pH

Form

Advantages

Drawbacks

Sodium dichloroisocyanurate (NaDCC)

6

Granules

Easy to ship Long shelf-life Does not clog pipes

Smell

High-test hypochlorite (HTH)

11

Granules

Easy to ship Long shelf-life Does not clog pipes

Explosive Clogs pipes

Stabilized sodium hypochlorite

11

Liquid

Can be local Does not clog pipes

Shorter shelf-life Difficult to ship

9-11

Liquid

Can be on-site Does not clog pipes

Shorter shelf-life Difficult to ship Quality control?

Non-stabilized sodium hypochlorite

421 422



423

Page 22 of 29

Table 3. Observed log removals and standard deviations.

Surface type

Soil load

Rec.

E. coli log removal

STD (E. coli)

Phi6 log removal

STD (Phi6)

Do nothing

>6.6

0.0

4.1

0.0

Wipe

>6.6

0.0

4.1

0.0

Cover

6.0

0.4

3.4

0.0

Wipe & cover

6.0

0.3

3.4

0.0

Do nothing

>6.8

0.0

5.7

0.0

Wipe

>6.8

0.0

5.7

0.0

Cover

>7.3

0.0

5.5

0.0

Wipe & cover

7.2

0.2

5.5

0.0

Do nothing

6.4

0.0

4.8

0.0

Wipe

>6.4

0.0

4.8

0.0

Cover

6.4

0.2

3.1

0.0

Wipe & cover

>6.5

0.0

3.1

0.0

Do nothing

>6.9

0.0

3.6

0.0

Wipe

6.8

0.2

3.3

0.4

Cover

>6.8

0.0

5.5

0.0

Wipe & cover

6.7

0.4

5.5

0.0

Do nothing

6.7

0.8

3.9

0.1

Wipe

6.7

0.9

3.8

0.0

Cover

6.3

0.9

3.4

0.0

Wipe & cover

6.1

1.0

3.4

0.0

Do nothing

6.0

1.1

5.4

0.0

Wipe

5.9

1.0

3.2

0.0

Cover

6.4

0.9

5.5

0.0

Wipe & cover

6.3

0.9

5.5

0.0

Without

Stainless steel

With

Without

Nitrile

With

Without

Heavy duty tarp

With

424 425


Page 23 of 29


426

GRAPHICS

427

TOC Art.

428 429 430 431

432 433

Figure 1. Testing matrix.



Page 24 of 29

434 435

Figure 2: Number of CFU E. coli per unit area detected after disinfection.

436 437 438 439

The points correspond to the geometric mean of triplicate samples. Error bars represent standard errors of the mean. “ND” stands for “not detected”, and the theoretical detection limit was < 1 CFU/cm2. The points indicated as too numerous to count (TNTC) correspond to cases were all triplicates were TNTC after disinfection.

440 441 442

443 444

Figure 3. Statistical results of E. coli testing.

445

NSD stands for “no significant difference” based on the Kruskal-Wallis test (p

Surface Cleaning and Disinfection: Efficacy Assessment of Four

Recommend Documents