Legionella Isolation Scheme

Summary of Results External Quality Assessment of Water Microbiology Legionella Isolation from Water Samples

Distribution Number: G97 Sample Numbers: G97A, G97B

Distribution Date: August 2015 Results Due: 11 September 2015 Report Date: 28 September 2015

Samples prepared and Morolake Adedeji quality control tested by: Mahin Khadjavi Thamayanthy Ramesh Aneta Stranc

Data analysed by: Manchari Rajkumar Nita Patel

Report compiled by: Manchari Rajkumar Nita Patel

Authorised by: Nita Patel

This report must not be reproduced without permission of the organisers.

Public Health England Food and Environmental Proficiency Testing Unit (FEPTU) 61 Colindale Avenue London NW9 5EQ Tel: +44 (0)20 8327 7119 Fax: +44 (0)20 8200 8264 Email: [email protected]

The data in FEPTU reports is confidential Page 1 of 11 Printed at 10:25 on Monday, 28 September, 2015 If you require further information on the scheme please refer to:

Scheme Guide: https://www.gov.uk/government/publications/food-and-water-proficiency-testing-schemes-scheme-guide Scoring Guide: https://www.gov.uk/government/publications/food-and-water-proficiency-testing-schemes-scoring-systems-and-statistics General guidance for z-scores: Participants' enumeration results are converted intoz-scores using the following formula:

x = participants' result expressed (expressed as a log 10 value)

Z = (x-X) X = assigned value (participants' consensus median (expressed as a log 10 value)) σ σ = the fixed standard deviation for the examination (calculated by FEPTU) The σ-value expresses the acceptable difference between the individual participant 's result and the participants' consensus median. The σ-value used for calculating z-scores for all parameters in the Legionella Scheme is 0.55. A guide to interpreting z-scores follows, although laboratories must interpret their scores in the context of their own laboratory situation. z = -1.99 to +1.99 satisfactory z = -2 to -2.99 or +2 to +2.99 questionable z= < -3.00 or > + 3.00 unsatisfactory It is usually recommended that z-scores exceeding +/-2.0 are investigated to establish the possible cause. As a general rule, PHE recommends that all questionable and unsatisfactory results are investigated.

FEPTU Quality Control: A minimum of 10 LENTICULE discs, selected randomly from the batch, are examined for Legionella spp. at the dilutions indicated on the request/report forms. The FEPTU results are determined using a method based on ISO 11731:1998: Water quality - Detection and enumeration of Legionella, and are included in the 'intended results' letters which provide guidance for participants regarding the assigned values. Intended results letters are posted on the website immediately after every distribution; electronic notification of their availability is sent to all participants.

If you experience difficulties with any of the examinations please refer to section 17.0 of the Scheme Guide https://www.gov.uk/government/publications/food-and-water-proficiency-testing-schemes-scheme-guide

All participants are reminded that reporting incorrect or false negative results for water samples could have serious public health implications.

Identification of Legionella spp. There are at least 50 species of Legionella of which at least 20 have been associated with human disease. The species that is most commonly isolated both from the environment and from infections is L.pneumophila; this species can be divided into at least 16 serogroups on the basis of surface antigens. L.pneumophila serogroup 1 is the most common cause of outbreaks of Legionnaires' disease and, within serogroup 1, the strains most commonly associated with disease share a common epitope, as revealed by monoclonal subtyping. It is important, because of the risks to human health, to differentiate routinely between L.pneumophila and other Legionella species and to be able to distinguish serogroup 1 from the other serogroups of L.pneumophila. However, monoclonal subtying is normally performed only by specialist reference laboratories. Participants who do not identity Legionella spp. other than L.pneumophila should where appropriate, report their results as 'Legionella spp. not L.pneumophila' rather than 'Legionella spp.'

Participants who report unexpectedly high or low results for the enumerations on single occasions only should not be unduly alarmed, although they should still assess the reason(s) for the outlying result. The results, as summarised in the performance assessment sheet included in this report, provide a more effective indication of on-going problems with Legionella examinations.

Please contact FEPTU staff for advice and information: Repeat samples Carmen Gomes or Kermin Daruwalla Tel: +44 (0)20 8327 7119 Data Analysis Manchari Rajkumar or Nita Patel Fax: +44 (0)20 8200 8264 Microbiological advice Morolake Adedeji or Nita Patel Email: [email protected] General comments and complaints Morolake Adedeji or Nita Patel Scheme consultants Dr John V Lee or Julie E. Russell Scheme Co-ordinator Nita Patel

Accreditation: PHE Water Scheme for Legionella Isolaton is accredited by the United Kingdom Accreditation Service (UKAS) to ISO/IEC Guide 17043:2010.

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The data in FEPTU reports is confidential Page 2 of 11 Printed at 10:25 on Monday, 28 September, 2015 Sample: G97A

Contents: l Legionella oakridgensis (wild strain) l Escherichia coli (wild strain)

Expected Results:

Expected Result Your Result PHE Score Z-Score Comments

Isolation of Detected legionellae

Identification L.oakridgensis

Serogroup N/A

Enumeration 5.3x10¹ - 9.13x10³ cfu L ⁻¹

Results Returned

Total score out of N/A

Background Flora: Escherichia coli formed colonies on GVPC medium after processing.

Comments on Performance:

Total participants reporting results 100

Participants reporting detected 52 (52%)

Participants reporting L.oakridgensis 11 (21%)

Participants reporting Legionella spp. not L.pneumophila 23 (44%)

Participants reporting Legionella spp. 10 (19%)

Participants reporting L.pneumophila 3 (6%)

Participants reporting not examined 1 (2%)

Serogroups for L.oakridgensis Participants reporting serogroup not applicable 41 (79%)

The data in FEPTU reports is confidential Page 3 of 11 Printed at 10:25 on Monday, 28 September, 2015 Sample: G97A (cont)

Total participants enumerating legionellae 99

Participants reporting a low censored value 44

Participants reporting a high censored value 1

Assigned value (participants' median) 1.00x10³ cfu L ⁻¹

Uncertainty of assigned value (Ux=log₁₀ cfu L⁻¹) 0.16 No.of outlying counts 12 (6 low / 6 high) Participants' mean 9.57x10² cfu L ⁻¹

Standard deviation of participants' results * 0.86 log₁₀ unit per cfu L ⁻¹

FEPTU median 3.30x10² cfu L ⁻¹

Total sent sample 114 Non-returns 7 Not examined 7

The fixed standard deviation value (σ value) used for calculation of the z-scores is 0.55 for all parameters. * Robust SD based on median absolute deviation about the participants' median (MAD).

The data in FEPTU reports is confidential Page 4 of 11 Printed at 10:25 on Monday, 28 September, 2015 Sample: G97B

Contents: l Legionella pneumophila (wild strain) l Enterobacter cloacae (wild strain) l Brevibacillus laterosporus (wild strain)

Expected Results:

Expected Result Your Result PHE Score Z-Score Comments

Isolation of Detected legionellae

Identification L.pneumophila

Serogroup 2 - 15

Enumeration 8.3x10¹ - 2.62x10³ cfu L ⁻¹

Results Returned

Total score out of N/A

Background Flora: Enterobacter cloacae formed colonies and Brevibacillus laterosporus did not form colonies on GVPC medium after processing.

Comments on Performance:

Total participants reporting results 101

Participants reporting detected 83 (82%)

Participants reporting L.pneumophila 75 (90%)

Participants reporting Legionella spp. not L.pneumophila 1 (1%)

Participants reporting not examined 1 (1%)

Participants reporting Legionella spp. 1 (1%)

Serogroups for L.pneumophila Participants reporting serogroups 2 - 15 76 (92%) Participants reporting no serogroup result 5 (6%) Participants reporting serogroup not applicable 1 (1%) Participants reporting serogroup not examined 1 (1%)

The data in FEPTU reports is confidential Page 5 of 11 Printed at 10:25 on Monday, 28 September, 2015 Sample: G97B (cont)

Total participants enumerating legionellae 100

Participants reporting a low censored value 15

Participants reporting a high censored value 1

Assigned value (participants' median) 4.67x10² cfu L ⁻¹

Uncertainty of assigned value (Ux=log₁₀ cfu L⁻¹) 0.07 No.of outlying counts 11 (5 low / 6 high) Participants' mean 4.58x10² cfu L ⁻¹

Standard deviation of participants' results * 0.49 log₁₀ unit per cfu L ⁻¹

FEPTU median 1.00x10² cfu L ⁻¹

Total sent sample 114 Non-returns 7 Not examined 6

The fixed standard deviation value (σ value) used for calculation of the z-scores is 0.55 for all parameters. * Robust SD based on median absolute deviation about the participants' median (MAD).

The data in FEPTU reports is confidential Page 6 of 11 Printed at 10:25 on Monday, 28 September, 2015 Performance Assessment Sheet

Participants are reminded that to take advantage of the performance assessment overtime tool provided in the reports they need to take part in more than one distribution a year.

Performance assessments are designed to identify laboratories with on -going problems with their examinations and are undertaken after every distribution. Scores are allocated to results reported for every sample to help assess participants ' performance.

Cumulative scores are calculated for every participant, for all examination types, for the current and previous three distributions. Participants' cummulative scores for each of the examination types are compared with the maximum possible scores after every distribution.

Distribution Sample Your score Your % Overall Performance Assessment for Distributions G97,G96,G95,G94 G97A G97 G97B 50 G96A G96 G96B 40 G95A G95 G95B 30 G94A G94 G94B 20 Number of participants

0 0 1-9 100 10-19 20-29 30-39 40-49 50-59 60-69 70-79 80-89 90-99 Percentage achieved of max possible score for stipulated distributions

Performance Assessment Comment: Laboratories that achieve less than 70% of the maximum possible score are likely to be experiencing significant problems with their examinations and are advised to a) refer to the relevant sample reports for specific comments b) refer to the website guidance documents: https://www.gov.uk/government/collections/external-quality-assessment-eqa-and-proficiency-testing-pt-for-food-water-and-environ mental-microbiology c) contact the organisers for advice.

The data in FEPTU reports is confidential Page 7 of 11 Printed at 10:25 on Monday, 28 September, 2015 Legionella counts reported by participants - Sample G97A

3 4 5 4 3 20 -1 3.00 Participant Median (log10 cfu L ) -1 2.52 FEPTU Median (log10 cfu L )

15 Outlying results Outlying results Outlying results Outlying results

10 Number of enumerations

0 <1.40 >4.34 1.40-1.72 1.73-1.83 1.84-1.93 1.94-2.03 2.04-2.13 2.14-2.24 2.25-2.34 2.35-2.44 2.45-2.54 2.55-2.64 2.65-2.74 2.75-2.84 2.85-2.95 2.96-3.05 3.06-3.15 3.16-3.25 3.26-3.35 3.36-3.45 3.46-3.55 3.56-3.66 3.67-3.76 3.77-3.96 3.96-4.34 cfu L ⁻¹ ( log₁₀)

Culture based presentation

G97A FEPTU Culture: GVPC

Culture Number Excluded Percentage Median Robust SD Range Reported of Results Results of the total Log 10 GVPC 44 34 81 3.00 0.73 1.38 - 4.04 BCYE with antibiotics 3 6 5 - BCYE agar without antibiotics 2 2 3 - PCR 1 1 1 - Other 4 2 7 -

G97A: Legionella counts reported by participants - Sample G97A 30 -1 3.00 Participant Median (log10 cfu L ) -1 2.52 FEPTU Median (log10 cfu L ) 25 Outlying results Outlying results Outlying results Outlying results 20

15 No. of Results

The data in FEPTU reports is confidential Page 8 of 11 Printed at 10:25 on Monday, 28 September, 2015 Legionella counts reported by participants - Sample G97B

3 4 5 4 3 30 -1 2.67 Participant Median (log10 cfu L ) -1 2.00 FEPTU Median (log10 cfu L )

25 Outlying results Outlying results Outlying results Outlying results 20

15 Number of enumerations 10

0 <1.72 >3.54 1.72-1.92 1.93-1.99 2.00-2.06 2.07-2.13 2.14-2.19 2.20-2.26 2.27-2.33 2.34-2.40 2.41-2.47 2.48-2.53 2.54-2.60 2.61-2.67 2.68-2.74 2.75-2.81 2.82-2.87 2.88-2.94 2.95-3.01 3.02-3.08 3.09-3.15 3.16-3.21 3.22-3.28 3.29-3.42 3.42-3.54 cfu L ⁻¹ ( log₁₀)

Culture based presentation

G97B FEPTU Culture: GVPC

Culture Number Excluded Percentage Median Robust SD Range Reported of Results Results of the total Log 10 GVPC 67 12 79 2.65 0.41 0.00 - 4.00 BCYE with antibiotics 7 2 8 - BCYE agar without antibiotics 3 1 3 - PCR 1 1 1 - Other 6 0 7 -

G97B: Legionella counts reported by participants - Sample G97B 40 -1 2.67 Participant Median (log10 cfu L ) -1 2.00 FEPTU Median (log10 cfu L ) 35

30 Outlying results Outlying results Outlying results Outlying results

20 No. of Results

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