Francesca Losito

Microbiological safety and quality of Mozzarella cheese assessed by the microbiological survey method

Dairy products are characterized by reduced shelf life because they are an excellent growth medium for a wide range of microorganisms. For this reason, it is important to monitor the microbiological quality of dairy products and, in particular, the total viable count and concentration of Escherichia coli, as they are indicators of the hygienic state of these products. In addition, in dairy products such as Mozzarella cheese, it is important to monitor the concentration of lactic acid bacteria (LAB), as they are the major components of starter cultures used in cheese production, contributing to the taste and texture of fermented products and inhibiting food spoilage bacteria by producing growth-inhibiting substances. For these reasons, to ensure the quality and safety of their products, cheese makers should monitor frequently, during fresh cheese production, the concentration of LAB and spoilage bacteria. However, usually, small- to medium-size dairy factories do not have an internal microbiological laboratory and external laboratories of analysis are often too expensive and require several days for the results. Compared with traditional methods, the microbiological survey (MBS) method developed by Roma Tre University (Rome, Italy) allows faster and less-expensive microbiological analyses to be conducted wherever they are necessary, without the need for a microbiological laboratory or any instrumentation other than MBS vials and a thermostat. In this paper, we report the primary validation of the MBS method to monitor LAB concentration in Mozzarella cheese and the analysis, using the MBS method, of total viable count, E. coli, and LAB concentrations in the production line of Mozzarella cheese as well as during the shelf life of the product stored at 20°C. The results obtained indicate that the MBS method may be successfully used by small- to medium-size dairy factories that do not have an internal microbiological laboratory. Using the MBS method, these dairy factories can monitor autonomously the microbiological safety and quality of their products, saving both time and money.

Field Application of the Micro Biological Survey Method for a Simple and Effective Assessment of the Microbiological Quality of Water Sources in Developing Countries

According to the World Health Organization (WHO) guidelines, “safe drinking-water must not represent any significant risk to health over a lifetime of consumption, including different sensitivities that may occur between life stages”. Traditional methods of water analysis are usually complex, time consuming and require an appropriately equipped laboratory, specialized personnel and expensive instrumentation. The aim of this work was to apply an alternative method, the Micro Biological Survey (MBS), to analyse for contaminants in drinking water. Preliminary experiments were carried out to demonstrate the linearity and accuracy of the MBS method and to verify the possibility of using the evaluation of total coliforms in 1 mL of water as a sufficient parameter to roughly though accurately determine water microbiological quality. The MBS method was then tested “on field” to assess the microbiological quality of water sources in the city of Douala (Cameroon, Central Africa). Analyses were performed on both dug and drilled wells in different periods of the year. Results confirm that the MBS method appears to be a valid and accurate method to evaluate the microbiological quality of many water sources and it can be of valuable aid in developing countries.

A new device for the prompt diagnosis of urinary tract infections.

Abstract Background: Urinary tract infections (UTIs) are among the most common infectious diseases. Results obtained from conventional microbiological analysis of urine and antibiotic susceptibility testing are available only after a few days, delaying precise diagnosis and appropriate therapy. Micro Biological Survey (MBS) srl (a spin-off of Roma Tre University, Rome, Italy) has developed and patented an automated colorimetric test for rapid bacterial counting. In a preliminary validation study it was demonstrated that the results obtained with the MBS method are equivalent to the results obtained with conventional culture-based microbiological analysis. Methods: In this study, sterile urine samples were artificially contaminated with bacterial species that are most frequently responsible for UTIs. The MBS method was used to evaluate the presence of bacteria and their sensitivity to some of the most commonly used antibiotics in UTIs. Results: The MBS method was able to detect in a few hours the presence or absence of bacteria at clinically significant concentrations (>105 CFU/mL), and to provide their susceptibility pattern to a limited panel of antibiotics. Discussion: The results obtained demonstrate that the MBS point-of-care testing (POCT) device could be developed into a valuable aid for the management of UTIs, possibly addressing more precise diagnosis and appropriate therapy.

A New Method For Microbiological Analysis That Could Be Used For Point-Of-Care Testing (POCT)

Standardized microbiological methods used in clinical analysis are based on traditional microbial enrichment on selective media, possibly followed by characterization of bacteria with molecular methods. These techniques present several difficulties, such as the subjectivity in the interpretation of genetic, biochemical or morphological tests and the possible interference of biological matrices, specially when low levels of contamination are present. In addition, standard- ized microbiological analyses are characterised by the high cost of the method, both in terms of labor and supplies, and above all, by the long time needed to obtain definitive results (from 3 to 7 days). These reasons have led to the develop- ment and refinement of microbiological POCTs which are now available for several microorganisms, even thought no mi- crobiological POCT was up to now developed for the count of total viable bacteria (TVC) in serum, urine or other bio- logical fluids. MBS srl (a spin-off of Roma Tre University, Rome, Italy) has developed and patented an alternative method for selective counting of bacteria, called Micro Biological Survey (MBS) method. The MBS method is based on colorimetric survey performed in mono-use disposable reaction vials in which samples can be inoculated without any preliminary treatment. The analyses can be carried out by untrained personnel and anywhere they are necessary, without the need for any other instrumentation than a thermostated optical reader that can automatically detect the colour change providing the number of bacteria present into the sample. The MBS method measures the catalytic activity of redox enzymes in the main meta- bolic pathways of bacteria, allowing an unequivocal correlation between the observed enzymatic activity and the number of viable cells present in the samples. The time required for a color change is inversely related to the log of bacterial con- centration; like an enzymatic reaction, the greater the number of bacteria, the faster the color change. The objective of this study was the primary validation, in accord with ISO 13843:2003 (Guidance on validation of micro- biological methods), of the quantitative Micro Biological Survey (MBS) method for Total Viable Count (TVC). Valida- tion aims to compare the results obtained with an alternative method, in this case the MBS method, with the results ob- tained with the reference method. To verify the equivalence between the two methods different parameters were analyzed: selectivity, linearity and accuracy. The validation has shown that the MBS method gives similar results and is in agree- ment with the reference methods. The MBS method could therefore represent a worthy aid in microbiological analysis as POCT device without replacing the analysis carried out with traditional methods which are very precise though often long and complex.

Integrating running water monitoring tools with the Micro Biological Survey (MBS) method to improve water quality assessment

Running water habitats are among the most altered aquatic systems by human activities driving an increase in the organic components and the associated bacterial load as well. To contribute in improving the monitoring activities in running waters, here we tested the validity of the new Micro Biological Survey (MBS) method to specifically assess the bacterial load in running waters focusing on Total Viable Counts (at 22°C and 37°C) and Escherichia coli (at 44°C) in order to propose a new prognostic tool for watercourses. MBS method is an alternative colorimetric method for counting bacterial load in water and food samples that is easy to use and leads to a reliable and simple interpretation of results, being also faster and less expensive than traditional methods. Then, we compared MBS with the traditionally used reference method for the bacterial load, and with the most used biotic index for Italian watercourses based on the benthic invertebrates: the Extended Biotic Index (EBI). The last comparison was performed to validate the use of MBS in biomonitoring activities since the benthic invertebrate multi-species assemblage (and then EBI) alter own structure mainly depending on the organic component variation. During the first part of the study, the assessment of both linearity (regressions among bacterial concentrations) and accuracy (significant correlation between a measured value and a value used as reference) confirmed the validity of the MBS method. Second, the linear regressions between the three investigated microbial parameters vs. both physical-chemical descriptors and EBI, revealed the usefulness of MBS as a valid tool for routine microbiological analyses involved in rapid and easy field monitoring activities. This represents the first attempt to evaluate the river microbial status by exploiting the innovative MBS on running waters to propose it as new valuable monitoring tool in the biomonitoring field.

Field application of the Micro Biological Survey method for the assessment of the microbiological safety of different water sources in Horn of Africa and the evaluation of the effectiveness of Moringa oleifera in drinking water purification

Water monitoring requires expensive instrumentations and skilled technicians. In developing Countries as Africa, the severe economic restrictions and lack of technology make water safety monitoring approaches applied in developed Countries, still not sustainable. The need to develop new methods that are suitable, affordable, and sustainable in the African context is urgent. The simple, economic and rapid Micro Biological Survey (MBS) method does not require an equipped laboratory nor special instruments and skilled technicians, but it can be very useful for routine water analysis. The aim of this work was the application of the MBS method to evaluate the microbiological safety of different water sources and the effectiveness of different drinking water treatments in the Horn of Africa. The obtained results have proved that this method could be very helpful to monitor water safety before and after various purification treatments, with the aim to control waterborne diseases especially in developing Countries, whose population is the most exposed to these diseases. In addition, it has been proved that Moringa oleifera water treatment is ineffective in decreasing bacterial load of Eritrea water samples.

A Pilot Clinical Trial on a New Point-of-care Test for the Diagnosis and Fast Management of Urinary Tract Infections in the Emergency Department

Background: Urinary tract infections (UTIs) are among the most common bacterial infections occurring in both community and health care setting. Laboratory diagnosis of UTIs is attained by conventional urine culture which identifies and quantifies infecting bacteria, followed by antibiotic susceptibility testing. This approach is time consuming and requires a considerable workload. A user-friendly, automated test for rapid bacterial quantification has been developed by MBS (a spin-off of Roma Tre University, Rome, Italy) as a point-of-care test (POCT) for UTIs. The aim of this study was to perform a preliminary clinical trial of the new MBS POCT for diagnosis and management of UTIs. Methods: A prospective diagnostic accuracy evaluation study was performed in collaboration with the Emergency Department of the Azienda Ospedaliera Sant’Andrea of Rome (Italy), on 122 patients with clinically suspected UTIs. Results of the MBS POCT were compared with those of the routine tests for urine culture and antibiotic susceptibility. Results: The MBS POCT, used in the Emergency Department, provided a UTI diagnosis in < 5 hours with very high accuracy, sensitivity and specificity. However, antibiotic susceptibility evaluation provided some false resistant results, when exceedingly high concentrations of bacteria were present in urines. Conclusion: The MBS POCT represents a valuable diagnostic tool for the detection of UTI, substantially saving time and assuring comparable quality of results, ultimately facilitating the successful management of infections. As for conventional antibiotic susceptibility tests, the bacterial inoculum is critical for significance of results.