A MALDI-TOF MS Approach for Mammalian, Human, and Formula Milks’ Profiling

Human milk composition is dynamic, and substitute formulae are intended to mimic its protein content. The purpose of this study was to investigate the potentiality of matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS), followed by multivariate data analyses as a tool to analyze the peptide profiles of mammalian, human, and formula milks. Breast milk samples from women at different lactation stages (2 (n = 5), 30 (n = 6), 60 (n = 5), and 90 (n = 4) days postpartum), and milk from donkeys (n = 4), cows (n = 4), buffaloes (n = 7), goats (n = 4), ewes (n = 5), and camels (n = 2) were collected. Different brands (n = 4) of infant formulae were also analyzed. Protein content (<30 kDa) was analyzed by MS, and data were exported for statistical elaborations. The mass spectra for each milk closely clustered together, whereas different milk samples resulted in well-separated mass spectra. Human samples formed a cluster in which colostrum constituted a well-defined subcluster. None of the milk formulae correlated with animal or human milk, although they were specifically characterized and correlated well with each other. These findings propose MALDI-TOF MS milk profiling as an analytical tool to discriminate, in a blinded way, different milk types. As each formula has a distinct specificity, shifting a baby from one to another formula implies a specific proteomic exposure. These profiles may assist in milk proteomics for easiness of use and minimization of costs, suggesting that the MALDI-TOF MS pipelines may be useful for not only milk adulteration assessments but also for the characterization of banked milk specimens in pediatric clinical settings.

Download Full-text

A MALDI-TOF MS Approach for Mammalian, Human, and Formula Milks’ Profiling

10.20944/preprints201808.0349.v1 ◽

2018 ◽

Author(s):

Laura Di Francesco ◽

Francesco Di Girolamo ◽

Maurizio Mennini ◽

Andrea Masotti ◽

Guglielmo Salvatori ◽

...

Keyword(s):

Human Milk ◽

Protein Content ◽

Low Cost ◽

Milk Composition ◽

Maldi Tof Ms ◽

Milk Samples ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Infant Formulae ◽

Tof Ms

Human milk composition is dynamic and substitute formulae are intended to mimic its protein content. The purpose of this study was to investigate the potentiality of matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) followed by multivariate data analyses as a tool to analyze peptide profiling of mammalian, human and formula milks. Breast milk samples from women at different lactation stages (2 (n = 5), 30 (n = 6), 60 (n = 5), and 90 (n = 4) days postpartum), and milk from donkeys (n = 4), cows (n = 4), buffaloes (n = 7), goats (n = 4), ewes (n = 5), and camels (n = 2) were collected. Different brands (n = 4) of infant formulae were also analyzed. Protein content (<30 kDa) was analyzed by MS and data were exported for statistical elaborations. Mass spectra for each milk closely clustered together, whereas different milk samples resulted well separated. Human samples formed a cluster in which colostrum constituted a well-defined subcluster. None of the milk formulae correlated with animal or human milk, although specifically characterized and well correlated each other. These findings propose MALDI-TOF MS milk profiling as an analytical tool to discriminate, in a blinded way, different milk types. As each formula has a distinct specificity, shifting a baby from one to another formula implies a specific proteomic exposure. These profiles may assist in milk proteomics for easiness of use and low cost consuming, suggesting that the MALDI-TOF MS pipelines may result useful for milk adulteration assessment but also for the characterization of banked milk specimens in paediatric clinical settings.

Download Full-text

A novel and rapid approach to yeast differentiation using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

Spectroscopy An International Journal ◽

10.1155/2003/849530 ◽

2003 ◽

Vol 17 (1) ◽

pp. 31-38 ◽

Cited By ~ 5

Author(s):

Richard E. Sherburn ◽

Richard O. Jenkins

Keyword(s):

Mass Spectrometry ◽

Mass Spectra ◽

Time Of Flight ◽

Laser Desorption ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Tof Ms ◽

Matrix Assisted Laser ◽

Matrix Assisted Laser Desorption

Matrix-assisted laser desorption/ionisation time of flight mass spectrometry (MALDI-TOF-MS) was investigated as a method for the rapid identification of yeast cells. Following pretreatment of yeast samples with a cell wall digesting enzyme (lyticase), distinct and reproducible mass spectra over them/zrange 2,000 to 16,000 were obtained by MALDI-TOF-MS. Using an optimised procedure, characteristic mass spectra that distinguished between Candida spp. and between strains of Saccharomyces cerevisiae were produced. The approach offers the potential for rapid differentiation of yeasts in clinical diagnosis and in the fermentation industries.

Download Full-text

Identification of Haemophilus influenzae Type b Isolates by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

Journal of Clinical Microbiology ◽

10.1128/jcm.00137-15 ◽

2015 ◽

Vol 53 (7) ◽

pp. 2215-2224 ◽

Cited By ~ 19

Author(s):

Viktor Månsson ◽

Fredrik Resman ◽

Markus Kostrzewa ◽

Bo Nilson ◽

Kristian Riesbeck

Keyword(s):

Mass Spectra ◽

Laser Desorption Ionization ◽

Type B ◽

Ionization Time ◽

Content Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Maldi Biotyper ◽

Tof Ms

Haemophilus influenzaetype b (Hib) is, in contrast to non-type bH. influenzae, associated with severe invasive disease, such as meningitis and epiglottitis, in small children. To date, accurateH. influenzaecapsule typing requires PCR, a time-consuming and cumbersome method. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) provides rapid bacterial diagnostics and is increasingly used in clinical microbiology laboratories. Here, MALDI-TOF MS was evaluated as a novel approach to separate Hib from otherH. influenzae. PCR-verified Hib and non-Hib reference isolates were selected based on genetic and spectral characteristics. Mass spectra of reference isolates were acquired and used to generate different classification algorithms for Hib/non-Hib differentiation using both ClinProTools and the MALDI Biotyper software. A test series of mass spectra from 33 Hib and 77 non-Hib isolates, all characterized by PCR, was used to evaluate the algorithms. Several algorithms yielded good results, but the two best were a ClinProTools model based on 22 separating peaks and subtyping main spectra (MSPs) using MALDI Biotyper. The ClinProTools model had a sensitivity of 100% and a specificity of 99%, and the results were 98% reproducible using a different MALDI-TOF MS instrument. The Biotyper subtyping MSPs had a sensitivity of 97%, a specificity of 100%, and 93% reproducibility. Our results suggest that it is possible to use MALDI-TOF MS to differentiate Hib from otherH. influenzae. This is a promising method for rapidly identifying Hib in unvaccinated populations and for the screening and surveillance of Hib carriage in vaccinated populations.

Download Full-text

Staphylococcus argenteus: An emerging subclinical bovine mastitis pathogen in Thailand

Veterinary World ◽

10.14202/vetworld.2019.1940-1944 ◽

2019 ◽

Vol 12 (12) ◽

pp. 1940-1944

Author(s):

Natapol Pumipuntu

Keyword(s):

Bovine Mastitis ◽

Bacterial Pathogen ◽

Technical Difficulty ◽

The Novel ◽

Ionization Time ◽

Maldi Tof Ms ◽

Milk Samples ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Tof Ms

Background and Aim: Staphylococcus argenteus is an emerging species of the Staphylococcus aureus complex. It has usually been misidentified as S. aureus by conventional methods and its characteristics. S. argenteus is potentially emerging in both humans and animals with an increasing global distribution. This study aimed to differentiate and identify S. argenteus from S. aureus collected and isolated from milk samples of subclinical bovine mastitis cases in Maha Sarakham Province, Northeastern Thailand. Materials and Methods: Forty-two isolates of S. aureus were studied from 132 individual milk samples collected from subclinical bovine mastitis cases of 15 dairy farms in three districts of Maha Sarakham, Thailand. The identification was confirmed by conventional and immune-agglutination methods. Fifteen representative isolates which were suspected as being S. argenteus were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results: The result from MALDI-TOF MS confirmed that seven from 15 isolates were S. argenteus and eight isolates were S. aureus. Conclusion: This study indicated that MALDI-TOF MS used as an identification and classification method could accurately differentiate the novel species, S. argenteus, from the S. aureus complex which is usually misdiagnosed. In addition, the identification of S. argenteus seems to be very limited in technical difficulty despite the fact that it may be the important causative pathogen in bovine mastitis as well as a pathogenic bacterium in food and milk. Therefore, it is essential for both bovine medicine and veterinary public health to emphasize and recognize this bacterial pathogen as an emerging disease of staphylococcal bacteria that there is a need for further study of S. argenteus infections.

Download Full-text

Test Agreement among Biochemical Methods, Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry, and 16S rRNA Sequencing for Identification of Microorganisms Isolated from Bovine Milk

Journal of Clinical Microbiology ◽

10.1128/jcm.01381-18 ◽

2018 ◽

Vol 57 (3) ◽

Cited By ~ 2

Author(s):

David J. Wilson ◽

John R. Middleton ◽

Pamela R. F. Adkins ◽

Gregory M. Goodell

Keyword(s):

16S Rrna ◽

Species Level ◽

Content Type ◽

Maldi Tof Ms ◽

Milk Samples ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Identification Of Bacteria ◽

Biochemical Testing ◽

Tof Ms

ABSTRACT The objective of this prospective study was a blinded comparison of three methods for the identification of bacteria isolated on Columbia blood agar from milk samples of dairy cows. Basic biochemical testing, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS), and 16S rRNA partial genome sequence analysis were compared for bacterial identification to the genus or species level. Milk samples submitted from a commercial dairy farm from recently calved cows or clinical mastitis cases were cultured, and 181 isolates were identified by biochemical testing, MALDI-TOF MS, and 16S rRNA sequence analysis (179 isolates; 2 isolates could not be recovered from storage). For Staphylococcus aureus and Escherichia coli, agreement was determined at the species level. For other microbes, agreement was determined at the genus level or at the group level for streptococcus-like organisms. The positive agreement among all 3 diagnostic methods was 94%, with 95% to 98% between each pair of methods. The overall (including negative agreement) agreement among all 3 methods ranged from 97% to 100%. MALDI-TOF MS is becoming more commonplace for the genus- and/or species-level identification of bacteria isolated from milk samples and, in some laboratories, has replaced conventional biochemical methods. The results of the present study suggest that when identifying pathogens at the genus or group level, conventional culture followed up with either secondary biochemical testing or MALDI-TOF MS is of practical value. For purposes of milk quality and udder health monitoring or research, any of the 3 methods is a valuable tool for genus-level identification of bacteria isolated from dairy cow milk.

Download Full-text

Identification of the ‘Streptococcus anginosus group’ by matrix-assisted laser desorption ionization – time-of-flight mass spectrometry

Journal of Medical Microbiology ◽

10.1099/jmm.0.076653-0 ◽

2014 ◽

Vol 63 (9) ◽

pp. 1143-1147 ◽

Cited By ~ 12

Author(s):

Katherine Woods ◽

David Beighton ◽

John L. Klein

Keyword(s):

Species Level ◽

Direct Transfer ◽

Laser Desorption Ionization ◽

Ionization Time ◽

Maldi Tof Ms ◽

Streptococcus Anginosus ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Transfer Method ◽

Tof Ms

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) provides rapid, accurate and cost-effective identification of a range of bacteria and is rapidly changing the face of routine diagnostic microbiology. However, certain groups of bacteria, for example streptococci (in particular viridans or non-haemolytic streptococci), are less reliably identified by this method. We studied the performance of MALDI-TOF MS for identification of the ‘Streptococcus anginosus group’ (SAG) to species level. In total, 116 stored bacteraemia isolates identified by conventional methods as belonging to the SAG were analysed by MALDI-TOF MS. Partial 16S rRNA gene sequencing, supplemented with sialidase activity testing, was performed on all isolates to provide ‘gold standard’ identification against which to compare MALDI-TOF MS performance. Overall, 100 % of isolates were correctly identified to the genus level and 93.1 % to the species level by MALDI-TOF MS. However, only 77.6 % were correctly identified to the genus level and 59.5 % to the species level by a MALDI-TOF MS direct transfer method alone. Use of a rapid in situ extraction method significantly improved identification rates when compared with the direct transfer method (P<0.001). We recommend routine use of this method to reduce the number of time-consuming full extractions required for identification of this group of bacteria by MALDI-TOF MS in the routine diagnostic laboratory. Only 22 % (1/9) of Streptococcus intermedius isolates were reliably identified by MALDI-TOF MS to the species level, even after full extraction. MALDI-TOF MS reliably identifies S. anginosus and Streptococcus constellatus to the species level but does not reliably identify S. intermedius.

Download Full-text

Rapid Classification of Clostridioides difficile Strains Using MALDI-TOF MS Peak-Based Assay in Comparison with PCR-Ribotyping

Microorganisms ◽

10.3390/microorganisms9030661 ◽

2021 ◽

Vol 9 (3) ◽

pp. 661

Author(s):

Adriana Calderaro ◽

Mirko Buttrini ◽

Monica Martinelli ◽

Benedetta Farina ◽

Tiziano Moro ◽

...

Keyword(s):

Ionization Time ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Clostridioides Difficile ◽

Pcr Ribotyping ◽

Typing Methods ◽

Set Up ◽

Tof Ms

Typing methods are needed for epidemiological tracking of new emerging and hypervirulent strains because of the growing incidence, severity and mortality of Clostridioides difficile infections (CDI). The aim of this study was the evaluation of a typing Matrix-Assisted Desorption/Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS (T-MALDI)) method for the rapid classification of the circulating C. difficile strains in comparison with polymerase chain reaction (PCR)-ribotyping results. Among 95 C. difficile strains, 10 ribotypes (PR1–PR10) were identified by PCR-ribotyping. In particular, 93.7% of the isolates (89/95) were grouped in five ribotypes (PR1–PR5). For T-MALDI, two classifying algorithm models (CAM) were tested: the first CAM involved all 10 ribotypes whereas the second one only the PR1–PR5 ribotypes. Better performance was obtained using the second CAM: recognition capability of 100%, cross-validation of 96.6% and agreement of 98.4% (60 correctly typed strains, limited to PR1–PR5 classification, out of 61 examined strains) with PCR-ribotyping results. T-MALDI seems to represent an alternative to PCR-ribotyping in terms of reproducibility, set up time and costs, as well as a useful tool in epidemiological investigation for the detection of C. difficile clusters (either among CAM included ribotypes or out-of-CAM ribotypes) involved in outbreaks.

Download Full-text

Matrix-assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) as a Reliable Tool to Identify Species of Catalase-negative Gram-positive Cocci not Belonging to the Streptococcus Genus

The Open Microbiology Journal ◽

10.2174/1874285801610010202 ◽

2016 ◽

Vol 10 (1) ◽

pp. 202-208 ◽

Cited By ~ 7

Author(s):

Marisa Almuzara ◽

Claudia Barberis ◽

Viviana Rojas Velázquez ◽

Maria Soledad Ramirez ◽

Angela Famiglietti ◽

...

Keyword(s):

Extraction Methods ◽

Species Level ◽

Ionization Time ◽

Maldi Tof Ms ◽

Genus Level ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Level Identification ◽

Gram Positive Cocci ◽

Tof Ms

Objective:To evaluate the performance of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) by using 190 Catalase-negative Gram-Positive Cocci (GPC) clinical isolates.Methods:All isolates were identified by conventional phenotypic tests following the proposed scheme by Ruoff and Christensen and MALDI-TOF MS (Bruker Daltonics, BD, Bremen, Germany). Two different extraction methods (direct transfer formic acid method on spot and ethanol formic acid extraction method) and different cut-offs for genus/specie level identification were used. The score cut-offs recommended by the manufacturer (≥ 2.000 for species-level, 1.700 to 1.999 for genus level and <1.700 no reliable identification) and lower cut-off scores (≥1.500 for genus level, ≥ 1.700 for species-level and score <1.500 no reliable identification) were considered for identification. A minimum difference of 10% between the top and next closest score was required for a different genus or species.MALDI-TOF MS identification was considered correct when the result obtained from MS database agreed with the phenotypic identification result.When both methods gave discordant results, the 16S rDNA orsodAgenes sequencing was considered as the gold standard identification method. The results obtained by MS concordant with genes sequencing, although discordant with conventional phenotyping, were considered correct. MS results discordant with 16S orsodA identification were considered incorrect.Results:Using the score cut-offs recommended by the manufacturer, 97.37% and 81.05% were correctly identified to genus and species level, respectively. On the other hand, using lower cut-off scores for identification, 97.89% and 94.21% isolates were correctly identified to genus and species level respectively by MALDI-TOF MS and no significant differences between the results obtained with two extraction methods were obtained.Conclusion:The results obtained suggest that MALDI-TOF MS has the potential of being an accurate tool for Catalase-negative GPC identification even for those species with difficult diagnosis asHelcococcus,Abiotrophia,Granulicatella, among others. Nevertheless, expansion of the library, especially including more strains with different spectra on the same species might overcome potential “intraspecies” variability problems. Moreover, a decrease of the identification scores for species and genus-level identification must be considered since it may improve the MALDI-TOF MS accuracy.

Download Full-text

Discrimination of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Journal of Veterinary Diagnostic Investigation ◽

10.1177/1040638717702687 ◽

2017 ◽

Vol 29 (5) ◽

pp. 622-627 ◽

Cited By ~ 7

Author(s):

Rinosh J. Mani ◽

Anil J. Thachil ◽

Akhilesh Ramachandran

Keyword(s):

Laser Desorption Ionization ◽

Biochemical System ◽

Ionization Time ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Subspecies Level ◽

Streptococcus Equi ◽

Level Identification ◽

Tof Ms

Accurate and timely identification of infectious etiologies is of great significance in veterinary microbiology, especially for critical diseases such as strangles, a highly contagious disease of horses caused by Streptococcus equi subsp. equi. We evaluated a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platform for use in species- and subspecies-level identification of S. equi isolates from horses and compared it with an automated biochemical system. We used 25 clinical isolates each of S. equi subsp. equi and S. equi subsp. zooepidemicus. Using the MALDI-TOF MS platform, it was possible to correctly identify all 50 isolates to the species level. Unique mass peaks were identified in the bacterial peptide mass spectra generated by MALDI-TOF MS, which can be used for accurate subspecies-level identification of S. equi. Mass peaks (mass/charge, m/ z) 6,751.9 ± 1.4 (mean ± standard deviation) and 5,958.1 ± 1.3 were found to be unique to S. equi subsp. equi and S. equi subsp. zooepidemicus, respectively. The automated biochemical system correctly identified 47 of 50 of the isolates to the species level as S. equi, whereas at the subspecies level, 24 of 25 S. equi subsp. equi isolates and 22 of 25 S. equi subsp. zooepidemicus isolates were correctly identified. Our results indicate that MALDI-TOF MS can be used for accurate species- and subspecies-level identification of S. equi.

Download Full-text