Fast detection of bacterial growth by using Portable Microbe Enrichment Unit (PMEU) and ChemPro100i® Gas Sensor.

The series of scientific articles, based on the microbiological research beneficing PMEU methods, continues. The new model of PMEU, "Scentrion", measures the gaseous metabolites, derived from the microbial activity in the sample syringes, and gives therefore the fastest indication of very low contamination levels in only 2..3 hours. PMEU Scentrion is already applied in a Finnish hospital laboratory. It is one of the new PMEU innovations, presented in a short R&D period in 2009 (another is PMEU "Spectrion" where the detection of the microbial growth bases on automatic turbidity measurement - more about it in future posts of this blog).

A short notification of this article follows below (to have more detailled view into the method and results derived with it, please refer the original article, published in June 2009 by Pathophysiology):

"E. Hakalehto, J. Pesola, A. Heitto, B. Bhanj Deo, K. Rissanen, U. Sankilampi, T. Humppi, H. Paakkanen.

Fast Detection of Bacterial Growth by using Portable Microbe Enrichment Unit (PMEU) and ChemPro100i® Gas Sensor.

Pathophysiology 16:1 (2009) 57-62.

PMEU Scentrion gives results in a few hours from the cultures, starting from only single cells per ml to a few hundreds of cells per ml. It is therefore a clear improvement to even PCR methods, which normally require an overnight pre-enrichment to be practically sensitive enough."

A novel training course of the PMEU enrichment and the microscopy of enriched samples in March, 2010.

The microbiological microscopy courses by FINN-FIBER Ltd. have been very popular ones over years. Participants of this annual course learn the basics of light and UV epifluorescence microscopy, isolate microbes from the samples and document the results with digital microphotography into the form of PowerPoint reports.

A novel course, containing PMEU enrichments in the beginning of this 3-day course, is under planning on these days. Four teachers - every one is a specialist on her/his area of microbiology - will combine their efforts to cover a wide range of bacteria and moulds as the subjects of the enrichments, isolations and microscopic examinations.

The course will be held in Jyväskylä, Central Finland, in March 2010. More information follows soon.

Rapid Microbiological Control of Paper Machines with PMEU Method

E.Hakalehto. Department of Biosciences, University of Kuopio, P.O.B. 1627, FI-70211 Kuopio, Finland, and Finnoflag Oy and Samplion Oy, Siilinjärvi, Finland (elias.hakalehto@pp.inet.fi)
L. Heitto. Environmental Research of Savo-Karjala Oy, FI-70150, Finland
J.V.Mentu. Samplion Oy, Siilinjärvi, Finland
H.Keitel. KK-Net Ky, FI-07900 Loviisa, Finland


PMEU incubations can be applied to rapid simulations of microbial growth in process waters and on the surfaces of test coupons as well as to testing of biocides against both waterborne and biofilm microorganisms. Relative efficiencies of alternative biocides can be tested with PMEU in original process samples during one working day. Recovery of the microbial population, treated with fast-acting biocides, can be detected after an incubation period of 24 hours.

When ATP Assay is included as a rapid measurement of microbial activity during the PMEU test periods, a fast, compact and realistic test procedure is ready to use for microbiological control of paper machine processes, without any need for microbiological laboratory facilities. Observation of biofilm production and detection of hazardous microbes can also be performed whenever additional analyses (Epifluorescence microscopy for biofilm testing, confirmation tests for hazardous microbes) are available.

- Published in "Paperi ja Puu" Paper and Pulp Magazine, Finland, Nov. 2009.

PMEU in PIRA Symposium, 14.-15.10.2009, in Barcelona.

Paper industry applications of PMEU method will be presented in PIRA Symposium, Barcelona, both in an oral presentation of Key Account Manager Juha V. Mentu and in the exhibition, located in the conference hotel:

Paper Chemistry Symposium 2009
13 - 15 October 2009

Paper Chemistry Symposium 2009 is IntertechPira's flagship offering for the paper industry. This two day conference provides a rare opportunity to draw upon key industry speakers expertise in paper chemistry, paper science and best practice examples in the forest products industry.

Benefits of PMEU method in raw material control, HACCP, biocide testing, biofilm detection and activated sludge investigations will be highlighted in this symposium.

Welcome to visit us in Barcelona!

Fast detection of bacterial growth by using Portable Microbe Enrichment Unit (PMEU) and ChemPro100i® gas sensor

Elias Hakalehto, Jouni Pesola, Anneli Heitto, Brajesh Bhanj Deob, Kari Rissanen, Ulla Sankilampi, Tarmo Humppi and Heikki Paakkanen

Abstract

The combination of the Portable Microbe Enrichment Unit (PMEU) cultivation and ChemPro100i® Chemical Detector helps the fast detection of hospital infections. The first alarm of the microbe metabolism and growth in the enrichment culture was achieved in 2–3 h when pure cultures of strains isolated from neonatal patients were tested. For the detection of minor concentrations of the bacteria, Bacillus, Staphylococcus, Klebsiella and Escherichia strains were diluted to 0.5–600 cells per ml. Their preliminary detection was achieved within 4–5 h. The combination of enrichment culture (PMEU) and detection of bacterial products by ion mobility spectrometer is a rapid and sensitive method for diagnosis of bacterial growth. This finding warrants further studies with clinical samples.

Keywords: Intestinal microbiota; Portable Microbe Enrichment Unit; Ion mobility spectrometer; Bacterial enrichment; Enterobacteriaceae; Escherichia coli; Klebsiella; Staphylococcus; Bacillus cereus; Hospital infections; Blood cultures; Neonatal septicaemia

The First PMEU Application: Rapid Detection of Harmful/Hazardous Bacteria.

The very first type of PMEU applications, selective detection of bacterial species, has been used worldwide in research institutes and hospitals since the end of 20th century. The schematic picture above describes the significant benefit of PMEU analyses, the speed. Selective cultivations can be started even at the sampling site and be continued during the transport of the samples to the receiving laboratory. Collection and pre-enrichment of samples for eg. environmental control (water authorities etc.) and HACCP quality management (food, dairy and beverage industry, packaging industry) are therefore available with highest speed found in microbiology today. The rapid detection of pathogens can even save human lifes in cases when the right antibiotic treatment of patients, suffering of infections in blood circulation system, can be found in time.

A new PMEU application: quantitative MPN analyses of microbial counts

PMEU method is based on the rapid cultivations of several samples. The old idea to apply it in MPN (Most Probable Number) analyses has now proven to be correct: referring the Finnish Standard Book "SFS-KÄSIKIRJA 94: Mikrobiologiset vesitutkimusmenetelmät" (Methods for Microbiological Water Analysis) and discussions with specialists, PMEU can be used as an alternative, rapid method instead of the traditional technique, tube series in water bath or in an incubator. PMEU itself works as an incubator with a temperature deviation of < 0.1 oC.

A combination of 4 (levels of dilutions) * 5 (repeats) allows to follow the Finnish standard SFS 4447 (The Tube Method in Microbiological Water Analysis) as well as standards derived of it like SFS-EN ISO 9308-3 (for and coliforms) and SFS-EN ISO 7899-1 (for enterococci). Standards usually give MPN tables in the framework of 3*5 tubes (eg. for dilutions from 0 to 0.01) but PMEU gives an extra level (eg. 0 to 0.001) which covers a wider range of microbial counts. Samples with unknown levels of microbial densities are therefore easier to analyse correctly.

It seems that the leading status of membrane filtration has revised today. There are types of samples which are difficult or impossible to analyse with them (too much suspended solids etc.) and tube tests like MPN should be chosen. PMEU Tube Tests should be preferred also in situations where fast results (in hours, compared with days with colony count analyses) are needed.

Applications of PMEU method for biofilm research and testing of biocides against biofilm growth

Paper machine biofilms have been studied already several years by microbiologist J.Mentu. Test coupons, made of steel brands used in paper machines, have been installed inside PMEU syringaes and the growth has been observed with UV Epifluorescence Microscopy after a short incubation period (see picture above).

This technique has now been modified for ordinary light microscopes, too. Steel coupons have been replaced by specified glass slides and the Gram-stained biofilms can be observed with Bright Field Microscopy - no expensive epifluorescence microscopes are needed in this application.

This method will detect all biofilm-producing microbes and testing of biofilm-preventing biocides is also possible simultaneously. Primary attachers typically appear on the slides in just hours and mature biofilms are available in 12...24 hours. This application is very suitable for all areas of industry where the hygiene of surfaces is important. It can also be applied in every environmental research projects where the formation of biofilms in natural water environments is the subject of the study. Hygiene control of public swimming pools etc. also benefit of this method.

World Record Incubator for Microbiological Research and Control Tasks

Enhanced enrichment and detection of thermotolerant Campylobacter species from water using the Portable Microbe Enrichment Unit and real-time PCR

Tarja Pitkänen, Juliane Bräcker, Ilkka T. Miettinen, Anneli Heitto, Jouni Pesola, and Elias Hakalehto

Can. J. Microbiol. 55(7): 849–858 (2009):

Abstract: An enhanced enrichment using the Portable Microbe Enrichment Unit (PMEU) with the microaerobic bubbling of broths was applied for the detection of thermotolerant Campylobacter species from water. This PMEU enrichment was compared with the conventional static enrichment of the international standard ISO 17995:2005. In addition, Campylobacter detection after enrichment using a real-time PCR detection was compared with colony counts. The tests with stressed Campylobacter jejuni cells in drinking water indicated that the PMEU enrichment yielded a significantly higher number of Campylobacter cells in the Bolton broth compared with the conventional static incubation. Application of the real-time PCR technique shortened the Campylobacter detection time. This combination of method modifications can be used for Campylobacter detection from water and adds methodological repertoire for the rapid survey and management of waterborne outbreaks.

Key words: Campylobacter, enrichment, Portable Microbe Enrichment Unit, real-time PCR, water.

PMEU-teknologian perusteet / Basic descriptions of PMEU technology.

PMEU -teknologia on tarkoitettu nopeuttamaan merkittävästi standardien mukaisia mikrobien viljelymenetelmiä. Samalla mikrobit, kuten erilaiset bakteerit, homeet ja hiivat, saadaan nopeasti rikastettua molekyylibiologisten tutkimusmenetelmien vaatimiin pitoisuuksiin. PMEU-menetelmää onkin menestyksellisesti hyödynnetty mm. PCR:n,immunoanalyysien ja ATP -mittausten esikäsittelymenetelmänä.

PMEU:n avulla saadaan mikrobien tutkimuksesta

1. herkempää
2. nopeampaa
3. luotettavampaa

kuin ennen; laboratorioissa, klinikoilla, tuotantolinjastoilla ja maastossa. Sen avulla näytteet saadaan tutkimuksen kohteiksi heti, ilman näytteen laatua heikentäviä viiveitä. Myös ympäristöstressistä kärsineet solut saadaan tehokkaasti elpymään.

FT, MMM Elias Hakalehto
Mikrobiologi, bioteknologi
International Postgraduate Diploma in Biotechnology (U.K.)
Bioteknisen mikrobianalytiikan dosentti
Kuopion yliopisto


PMEU (Portable Microbe Enrichment Unit) technology has been designed tomake the standard microbe cultivation methods significantly faster. At the same time the microbes, such as bacteria, moulds and yeasts, are rapidly enriched to concentrations required for different molecular methods. PMEU method has been successfully used for pretreatments for the PCR,
immunoanalysis and ATP measurements.

PMEU approach makes the microbiological research

1. more sensitive
2. faster
3. more reliable

than before; in the laboratories, clinics, production lines and outdoors.
The samples are studies immediately after sampling without delays that
lower the sample quality. Also the cells suffered from environmental
stresses are effectively recovered.

Elias Hakalehto, PhD, MSc (Agr.Sci)
Microbiologist, Biotechnologist
International Postgraduate Diploma in Biotechnology (U.K.)
Docent in Biotechnical Microbe Analytics
University of Kuopio
Kuopio, Finland

PMEU applications for paper industry microbiology.

Paper industry microbiology is one of the several sectors of microbiology which can have significant benefits when applying PMEU methods for rapid hygiene control and HACCP procedures.

A paper mill consists of several technical parts which all act as growth environments (niches) for microbes. Paper machine itself is like a continuous flow fermentor, securing constant nutrient supply, growth environment and feedback for microbes. All regions of the system have their own microbial populations which cause risks for the processes, machinery, employees and product hygiene. Portable Microbiological Enrichment Unit (PMEU) by SAMPLION Oy (Siilinjärvi, Finland) has been modified to an analytical tool for paper industry microbiology, intended in estimation of microbial activities in process waters and simultaneous evaluation of alternative paper machine biocides. ATP Assay was combined with PMEU incubations to give quantitative values of living biomass during tests. PMEU can be used for the detection of harmful bacteria in raw materials and processes as well as on the control of biofilm-forming microbes when epifluorescence microscopy for the observations of biofilm growth on test plates is available. PMEU method helps to avoid bias, caused by the selecting and recovering effects of traditional colony count methods. Because of the robustness of  PMEU incubator, it does not necessarily need any specified microbiological laboratory but acts also as a stand alone cultivation system in common paper mill laboratories research institutes as their additional microbiological tool.

Marketing of PMEU unit is starting in this summer.

PMEU (Portable Microbiological Enrichment Unit) was developed by Dr. Elias Hakalehto for research purposes on 90's. A novel company called SAMPLION Ltd., grounded in May 2009, starts to deliver PMEU applications for different areas of microbiology.

Main features of PMEU enrichment, assay and testing procedures are its capability to enhance the growth of target microbes, its sensitivity and the challenge it serves when simulations of real target environments are needed. These features have already been proven in several research projects, derived eg. in hospitals, environmental laboratories and paper industry institutes.

We shall present these applications and refer our scientific articles, based on clinical, environmental, biotechnical and paper industry research projects, in this blog. All writers are experienced PMEU users, having long careers in microbiological research.

PORTABLE MICROBIOLOGICAL ENRICHMENT UNIT (PMEU)