Table 1 Overview of publications on cleaning intervention trials included in this narrative review. CFU, colony forming units., PCHS, Probiotic Cleaning Hygiene System. spp., subspecies

1

Vandini, Plos One, 2014 [38]

Probiotic Cleaning Hygiene System (PCHS; Copma srl, Italy), by using detergents containing 10⁷ spores per ml of B. subtilis, B. pumilus and B. megaterium (Chrisal, Lommel, Belgium)

24 weeks

Conventional cleaning:

chemical detergents in 1 Belgian hospital (Ecolab, Groot-Bijgaarden, Belgium),

In both Italian hospitals: chlorine-based detergents (Actichlor for all washable surfaces, Diversey S.p.A., Italy)

3 hospitals, Belgium (n = 1), Italy (n = 2)

Multicentre, pre-post interventional study

20,000 environmental samples

Presence and survival of several microorganisms responsible for HAIs (i.e. coliforms, Staphylococcus aureus, Clostridiodes difficile, and Candida albicans) on hard surfaces by CFU / m²

Reduction of HAI-related pathogens by 50 to 89%

2

Caselli, Plos One, 2016 [22]

See publication no. 1

6 months

No control group

1 hospital, Italy

Single centre, pre-post interventional study

360 environmental samples collected in duplicates (n = 720)

Growth of Staphylococcus spp. and Staphylococcus aureus, Enterobacterales, Acinetobacter spp., Pseudomonas spp., C. difficile, Candida spp. and Aspergillus spp. by CFU or genome copies/ m²;

Spore germination (in vitro),

antibiotic resistance genes

Decrease of the number of CFU/m² for all pathogens tested except for Enterobacterales due to scarce presence at T0,

Germination of Bacillus spores on dry inanimate surfaces demonstrated (in vitro),

Decrease of antibiotic resistance genes in the contaminating microbial population

3

Caselli, Plos One, 2018 [35]

See publication no. 1

6 months

Conventional chemical-based chlorine products

6 hospitals, Italy

Multicentre, pre-post interventional study

11,842 patients and 24,875 environmental samples

Qualitative and quantitative characterisation of hospital surface bioburden

Significant decrease of HAI cumulative incidence from a global 4.8% (284 patients with HAI over 5,930 total patients) to 2.3% (128 patients with HAI over 5,531 total patients) (OR = 0.44, CI 95% 0.35 ± 0.54) (P < 0.0001),

2 Log drop of surface microbiota drug-resistance

genes (P < 0.0001; Pc = 0.008)

4

Caselli, Infection and Drug Resistance, 2019 [34]

Additional analysis of trial no.3 [22]

6 months

Conventional chemical-based chlorine products

5 hospitals, Italy

Multicentre, pre-post interventional study

756 environmental samples,

antimicrobial drug consumption analysis and economic analysis based on 398 patients with HAI and data available

Surface microbiota, AMR (by microarray, nested PCR, antibiogram, and microdilution tests),

antibiotic consumption and associated costs

Up to 99% decrease of the AMR genes harbored by surface hospital microbiota,

Antimicrobial drug consumption associated with HAI onset showed a global 60.3% decrease, with a 75.4% decrease of the associated costs.

5

Tarricone, Pathogens 2020 [33]

Economic analysis of trial no.3 [22], conventional chemical cleaning (CCC) versus PCHS

6 months

Conventional chemical-based chlorine products

6 hospitals, Italy

Economic analysis of trial no.3 [22]: Multicentre, pre-post interventional study with subsequent propensity-score matching (estimated increase of PCHS use from 5–50% in the next 5 years),

11,461 patients (propensity score matching)

Number of HAIs per 1000 patient days in both CCC and PCHS periods;

HAIs outcomes;

Consumption of drugs to treat HAIs and the related costs;

Identification of cases of antibiotic-resistance and estimation of their treatment costs

Cumulative incidence of HAI: 4.6% and 2.4% (p < 0.0001) for CCC (N = 4160) and PCHS (N = 4160), (OR = 0.47, CI 95% 0.37–0.60), with severe HAIs of 1.57% vs. 1% and antibiotic resistances of 1.13% vs. 0.53%,

Estimated savings by PCHS:

about 31,000 HAIs and 8,500 antibiotic resistances,

14 million euros, of which 11.6 for the treatment of resistant HAIs

6

Soffritti, Infect Drug Resist 2022 [37]

See publication no. 1

2 months

Conventional chemical-based chlorine products

1 hospital (Maternal and Child Health Institute), Italy

Single centre, pre-post interventional study

152 samples environmental samples

Microbial contamination (monitored by culture-based CFU count, 16 S rRNA NGS for bacteriome characterization and microarrays for resistome assessment),

SARS-CoV-2 presence (monitored by PCR)

PCHS usage was associated with a stable 80% decrease in surface pathogens compared to levels detected for chemical disinfection (P < 0.01),

up to 2 log decrease in resistance genes (P < 0.01)

SARS-CoV-2 was not detectable in the pre-PCHS and PCHS period.

6

D’Accolti Microbiome 2023 [46]

PCHS (prepreg cloths/ nebulization) was applied 30 min after ethanol/ammonium disinfection (see publication no. 1) versus chemical disinfection with chlorine-, ethanol-, and ammonium salt-based products

12 weeks

Routine chemical disinfection with chlorine-, ethanol-,

and ammonium salt-based products

Two underground driverless trains with four compartment each corresponding to a total of 96 seats and 438 standing places for a total 50.5 m in length and 73 m² of surface. 1 train with PCHS, 1 train with standard disinfection

Non-randomized, controlled interventional trial

6 sampling campaigns, sampling of 12 total points in duplicates from surfaces (floor, seats, handrails, doors) and air filters (if available), collected by RODAC plates /sterile swabs

Microbial growth (culture-based CFU count),

16SrDNA NGS, real-time qPCR microarray, NGS profiling

Clear and significant decrease in bacterial and fungal pathogens (p < 0.001), decrease of SARS-CoV-2 presence (p < 0.01) in the probiotic-treated train compared with the chemically disinfected control train

7

Klassert Clin Microbiol Infect 2022 [28]

0.5% detergents with concentration containing 10⁷ spores per ml B.  subtilis (ATCC6051), B.  megaterium (ATCC14581), B. licheniformis (ATCC12713), B. pumilus (ATCC14884) ,and B. amyloliquefaciens (DSL13563-0) (by Chrisal)

3 months

Soap-based arm: agent with non-ionic surfactants, anionic surfactants, and fragrances in a total concentration of 1% (Brial Top^®, Ecolab Inc.).

Disinfectant arm: 2-phenoxyethanol (10%), 3-aminopropyldodecylamine (8%), benzalkonium chloride (7.5%) at a total concentration of 1.5%, with a contact time of 15 min (Incidin Pro^®, Ecolab Inc.)

1 hospital, Germany

Single centre, pre-post interventional study

1,019 environmental samples for analysis of environmental microbiota

551 environmental samples for ARG detection

Environmental microbiota (by 16 S rRNA sequencing),

Detection of antibiotic resistance genes (ARGs) by multiplex Taq-Man qPCR assays

Significant increase in the alpha-diversity metrics samples from the floor (p < 0.001) and the sink (p < 0.01), but not from door handles during the probiotic strategy (compared with disinfectant),

Displacement of the intrinsic environmental microbiota in sink samples (median 16 S-rRNA copies = 138.3; IQR: 24.38–379.5) compared to traditional disinfection measures (median 16 S rRNA copies = 1343; IQR: 330.9–9479; p < 0.05).

Significant reduction in the total ARG counts in the sink samples during probiotic cleaning (mean ARGs/sample: 0.095 ± 0.067) compared to the disinfection strategy (mean ARGs/sample: 0.386 ± 0.116; p < 0.01)

8

Leistner, eClinicalMedicine, 2023 [32]

0.5% detergents containing 10⁷ spores per ml B. subtilis (ATCC6051), megaterium (ATCC14581), B. licheniformis (ATCC12713), B. pumilus (ATCC14884), and. amyloliquefaciens (DSL13563-0) (by Chrisal)

4 months

Soap-based arm: agent with non-ionic surfactants, anionic surfactants, and fragrances in a total concentration of 1% (Brial Top^®, Ecolab Inc.).

Disinfectant arm: 2-phenoxyethanol (10%), 3-aminopropyldodecylamine (8%), benzalkonium chloride (7.5%) at a total concentration of 1.5%, with a contact time of 15 min (Incidin Pro^®, Ecolab Inc.)

1 hospital, 18 wards, Germany

cRCT with cross-over design

13.896 patients from 18 wards

Incidence density of HAI

Incidence density of HAI with MDRO

Incidence density of HAI was similar between the reference group (soap), disinfectant and probiotics:

2.31 cases per 1000 exposure days for soap, 2.21 for disinfectant (IRR 0.95; 95% CI 0.69–1.31; p = 0.953) and 2.21 for probiotics (IRR 0.96; 95% CI 0.69–1.32; p = 0.955).

Incidence density of MDRO infection was similar between the reference group (soap), disinfectant and probiotics:

0.53 cases per 1000 exposure days (0.32–0.84) for soap versus 0.49 (0.29–0.78) for disinfectant and versus 0.46 (0.26–0.76) for probiotics

9

Dancer, BMC Med 2009 [52]

Enhanced cleaning with CINCH™ detergent (AGMA, Haltwhistle, UK) and water and Tuffie™ wipes (Health Care Services, Nottinghamshire, UK) by an additional cleaner

6 months per phase

Conventional cleaning with the same detergent and wipes

1 hospital; 2 wards, UK

Single-center, prospective cross-over trial

Environmental samples from 10 hand-touch sites (e.g. infusion pump, blood pressure, and computer keyboard) on both wards every week for 1 year

Microbial contamination on hand-touched sites,

Incidence MRSA infections,

Cost savings

Reduction of microbial contamination at hand-touched sites by 32.5%,

Reduced the number of new MRSA infections by 26.6%,

Cost savings between 30.000 and 70.000 pounds estimated