Relationships among intramammary health, udder and teat characteristics, and productivity of extensively managed ewes

Abstract Mastitis is an economically important disease and its subclinical state is difficult to diagnose, which makes mitigation more challenging. The objectives of this study were to screen clinically healthy ewes in order to 1) identify cultivable microbial species in milk, 2) evaluate somatic cell count (SCC) thresholds associated with intramammary infection, and 3) estimate relationships between udder and teat morphometric traits, SCC, and ewe productivity. Milk was collected from two flocks in early (<5 d) and peak (30 to 45 d) lactation to quantify SCC (n = 530) and numerate cultivable microbial species by culture-based isolation followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS; n = 243) identification. Within flock and lactation stage, 11% to 74% (mean = 36%) of samples were culture positive. More than 50 unique identifications were classified by MALDI-TOF MS analysis, and Bacillus licheniformis (18% to 27%), Micrococcus flavus (25%), Bacillus amyloliquefaciens (7% to 18%), and Staphylococcus epidermidis (26%) were among the most common within flock and across lactation stage. Optimum SCC thresholds to identify culture-positive samples ranged from 175 × 103 to 1,675 × 103 cells/mL. Ewe productivity was assessed as total 120-d adjusted litter weight (LW120) and analyzed within flock with breed, parity, year, and the linear covariate of log10 SCC (LSCC) at early or peak lactation. Although dependent on lactation stage and year, each 1-unit increase in LSCC (e.g., an increase in SCC from 100 × 103 to 1,000 × 103 cells/mL) was predicted to decrease LW120 between 9.5 and 16.1 kg when significant. Udder and teat traits included udder circumference, teat length, teat placement, and degree of separation of the udder halves. Correlations between traits were generally low to moderate within and across lactation stage and most were not consistently predictive of ewe LSCC. Overall, the frequencies of bacteria-positive milk samples indicated that subclinical mastitis (SCM) is common in these flocks and can impact ewe productivity. Therefore, future research is warranted to investigate pathways and timing of microbial invasion, genomic regions associated with susceptibility, and husbandry to mitigate the impact of SCM in extensively managed ewes.

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Optimized Preparation of Urine Samples from Acute Melioidosis Patients for In-Solution Proteomic Studies using LCMS QTOF or MALDI TOF MS

IIUM Medical Journal Malaysia ◽

10.31436/imjm.v20i3.1844 ◽

2021 ◽

Vol 20 (3) ◽

Author(s):

Aniza Pakeer ◽

Mohammed Imad A. Mustafa Mahmud ◽

Nosrihah Ismail ◽

Vanitha Mariappan

Keyword(s):

Biomarker Discovery ◽

Acute Phase Proteins ◽

Protein Precipitation ◽

Urine Samples ◽

Precipitation Method ◽

Cell Mediated Immunity ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Culture Positive ◽

Tof Ms

Introduction: Investigation of urine proteome in patients with acute melioidosis may reveal potential disease markers, from either bacterial or human proteins. We used an in-solution gel-free method instead of 2-DE to detect human and Burkholderia pseudomallei proteins in urine of patients with acute melioidosis. Here, we propose a simpler, economical method for preparing urine samples directly from melioidosis patients, for in-solution proteomic analysis using LCMS-QTOF MS/MS or MALDI-TOF MS/MS. Material and Methods: We adapted an acetone-TCA based protein precipitation method with LCMS-QTOF MS to detect the B. pseudomallei proteins directly from urine of acute melioidosis patients (culture positive and negative). This process involves protein precipitation, desalting, trypsin digestion, and optimization for the mass spectrometry. Results: A total of 3,866 human peptides were detected across 11 urine samples from clinically suspected acute melioidosis patients. Among them were three Burkholderia specific proteins detected in 75% of culture positive samples. Large amounts of acute phase proteins, cell mediated immunity proteins, complement pathway proteins and inflammatory mediators were seen upon gene ontology (GO) annotation and GO enrichment analysis. Conclusions: This simple in-solution sample preparation method can be replicated easily for LCMS/MS-QTOF and MALDI-TOF proteomic analyses, avoiding tedious optimization steps in 2-DE. This method is cost effective and can be done in centres without specialized 2-DE or MS equipment and elutes can be easily transported for analysis and bioinformatics. This is the first study to analyse urine samples directly for B. pseudomallei proteins. Discovery of the entire proteome as a whole is important in leading to biomarker discovery.

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Conserved mass peaks in MALDI-TOF mass spectra of bacterial species

10.7287/peerj.preprints.3524v1 ◽

2018 ◽

Author(s):

Wenfa Ng

Keyword(s):

Mass Spectrum ◽

Mass Spectra ◽

Bacterial Species ◽

Biological Basis ◽

Maldi Tof Ms ◽

Microbial Identification ◽

Microbial Species ◽

Maldi Tof ◽

Maldi Tof Mass Spectra ◽

Tof Ms

Microbes are identified based on their distinguishing characteristics such as gene sequence or metabolic profile. Nucleic acid approaches such as 16S rRNA gene sequencing provide the gold standard method for microbial identification in the contemporary era. However, mass spectrometry-based microbial identification is gaining credence through ease of use, speed, and reliability. Specifically, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used in identifying bacteria, fungus, molds and archaea to the species level with high accuracy. The approach relies on the existence of unique mass spectrum fingerprint for individual microbial species. By comparing the mass spectrum of an unknown microbe with that catalogued in a reference database of known microorganisms, microbes could be identified through mass spectrum fingerprinting. However, the approach lacks fundamental biological basis given the relative difficulty in assigning specific protein to particular mass peak in the profiled mass spectrum, which hampers a deeper understanding of the mass spectrum obtained. This study seeks to examine the existence of conserved mass peaks in MALDI-TOF mass spectra of bacterial strains belonging to the same species in open access data from SpectraBank. Results revealed that conserved mass peaks existed for all bacterial species examined (Bacillus subtilis, Bacillus thuringiensis, Carnobacterium maltaaromaticum, Escherichia coli, Proteus vulgaris, Pseudomonas fluorescens, Pseudomonas fragi, Pseudomonas putida, Pseudomonas syringae, Serratia marcescens, Serratia proteamaculans, Staphylococcus aureus, and Stenotrophomonas maltophilia). Large number of conserved mass peaks such as that of E. coli might suggest more closely-related strains of a species though functional annotation of the mass peaks is required to provide deeper understanding of the mechanisms underlying the conservation of specific proteins. On the other hand, strains of S. aureus and P. putida had the least number of conserved mass peaks. Presence of conserved mass peaks in the genus Pseudomonas and Serratia provided further evidence that MALDI-TOF MS microbial identification had a biological basis in identification of microbial species to the genus level. On the other hand, it also highlighted that a subset of proteins could define the taxonomical boundary between the species and genus level. Overall, existence of conserved mass peaks in strains of the same bacterial species provided evidence of a firm biological basis in the mass spectrum fingerprinting approach of MALDI-TOF MS microbial identification. This could help identify specific species in mass spectrum of single or multiple microbial species. Further functional annotation of the conserved mass peaks could illuminate in greater detail the biological mysteries of why certain proteins are conserved in specific genus and species.

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A new culture-based method for rapid identification of microorganisms in polymicrobial blood cultures by MALDI-TOF MS

BMC Microbiology ◽

10.1186/s12866-019-1641-1 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 5

Author(s):

Walter Florio ◽

Susanna Cappellini ◽

Cesira Giordano ◽

Alessandra Vecchione ◽

Emilia Ghelardi ◽

...

Keyword(s):

Rapid Identification ◽

Blood Cultures ◽

Time Saving ◽

Simple Method ◽

Maldi Tof Ms ◽

Microbial Identification ◽

Microbial Species ◽

Maldi Tof ◽

Identification Of Bacteria ◽

Tof Ms

Abstract Background The application of matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry (MS) to microbial identification has allowed the development of rapid methods for identification of microorganisms directly in positive, blood cultures (BCs). These methods can yield accurate results for monomicrobial BCs, but often fail to identify multiple microorganisms in polymicrobial BCs. The present study was aimed at establishing a rapid and simple method for identification of bacteria and yeast in polymicrobial BCs from patients with bloodstream infection. Results The rapid method herein proposed is based on short-term culture in liquid media allowing selective growth of microorganisms recovered from polymicrobial BCs, followed by rapid identification by MALDI-TOF MS. To evaluate the accuracy of this method, 56 polymicrobial BCs were comparatively analyzed with the rapid and routine methods. The results showed concordant identification for both microbial species in 43/50 (86%) BCs containing two different microorganisms, and for two microbial species in six BCs containing more than two different species. Overall, 102/119 (85.7%) microorganisms were concordantly identified by the rapid and routine methods using a cut-off value of 1.700 for valid identification. The mean time to identification after BC positivity was about 4.2 h for streptococci/enterococci, 8.7 h for staphylococci, 11.1 h for Gram-negative bacteria, and 14.4 h for yeast, allowing a significant time saving compared to the routine method. Conclusions The proposed method allowed rapid and reliable microbial identification in polymicrobial BCs, and could provide clinicians with timely, useful information to streamline empirical antimicrobial therapy in critically ill patients.

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A transient tobacco expression system coupled to MALDI-TOF-MS allows validation of the impact of differential targeting on structure and activity of a recombinant therapeutic glycoprotein produced in plants

FEBS Letters ◽

10.1016/s0014-5793(03)00916-5 ◽

2003 ◽

Vol 552 (2-3) ◽

pp. 170-176 ◽

Cited By ~ 11

Author(s):

Nathalie Mokrzycki-Issartel ◽

Bernadette Bouchon ◽

Sibille Farrer ◽

Patricia Berland ◽

Hélène Laparra ◽

...

Keyword(s):

Expression System ◽

Maldi Tof Ms ◽

Maldi Tof ◽

The Impact ◽

Structure And Activity ◽

Tof Ms

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The impact of delayed analysis of positive blood cultures on the performance of short-term culture followed by MALDI-TOF MS

Journal of Microbiological Methods ◽

10.1016/j.mimet.2020.106027 ◽

2020 ◽

Vol 177 ◽

pp. 106027

Author(s):

Alexander T.A. Johnsson ◽

Alicia Y.W. Wong ◽

Volkan Özenci

Keyword(s):

Blood Cultures ◽

Short Term ◽

Maldi Tof Ms ◽

Maldi Tof ◽

The Impact ◽

Short Term Culture ◽

Tof Ms

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The impact of blood culture identification by MALDI-TOF MS on the antimicrobial management of pediatric patients

Diagnostic Microbiology and Infectious Disease ◽

10.1016/j.diagmicrobio.2018.05.021 ◽

2018 ◽

Vol 92 (3) ◽

pp. 220-225 ◽

Cited By ~ 6

Author(s):

Sejal Makvana Bhavsar ◽

Tanis C. Dingle ◽

Camille L. Hamula

Keyword(s):

Blood Culture ◽

Pediatric Patients ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Antimicrobial Management ◽

Culture Identification ◽

The Impact ◽

Tof Ms

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Study on Molecular Profiles of Staphylococcus aureus Strains: Spectrometric Approach

Molecules ◽

10.3390/molecules25214894 ◽

2020 ◽

Vol 25 (21) ◽

pp. 4894

Author(s):

Michał Złoch ◽

Paweł Pomastowski ◽

Ewelina Maślak ◽

Fernanda Monedeiro ◽

Bogusław Buszewski

Keyword(s):

Staphylococcus Aureus ◽

Culture Conditions ◽

Strain Typing ◽

Major Health Problem ◽

Matrix Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Molecular Profiles ◽

The Impact ◽

Tof Ms

Staphylococcus aureus remains a major health problem responsible for many epidemic outbreaks. Therefore, the development of efficient and rapid methods for studying molecular profiles of S. aureus strains for its further typing is in high demand. Among many techniques, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI TOF MS) represents a timely, cost-effective, and reliable strain typing approach, which is still rarely used due to insufficient knowledge about the impact of sample preparation and analysis conditions on the molecular profiles and strain classification efficiency of S. aureus. The aim of this study was to evaluate the effect of the culture conditions and matrix type on the differentiation of molecular profiles of various S. aureus strains via the MALDI TOF MS analysis and different computational methods. The analysis revealed that by changing the culture conditions, matrix type, as well as a statistical method, the differentiation of S. aureus strains can be significantly improved. Therefore, to accelerate the incorporation of the MALDI-based strain typing in routine laboratories, further studies on the standardization and searching of optimal conditions on a larger number of isolates and bacterial species are of great need.

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Progress in proteomics for clinical microbiology: MALDI-TOF MS for microbial species identification and more

Expert Review of Proteomics ◽

10.1586/14789450.2015.1091731 ◽

2015 ◽

Vol 12 (6) ◽

pp. 595-605 ◽

Cited By ~ 47

Author(s):

Alex van Belkum ◽

Sonia Chatellier ◽

Victoria Girard ◽

David Pincus ◽

Parampal Deol ◽

...

Keyword(s):

Species Identification ◽

Clinical Microbiology ◽

Maldi Tof Ms ◽

Microbial Species ◽

Maldi Tof ◽

Tof Ms

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Review of the impact of MALDI-TOF MS in public health and hospital hygiene, 2018

Eurosurveillance ◽

10.2807/1560-7917.es.2019.24.4.1800193 ◽

2019 ◽

Vol 24 (4) ◽

Cited By ~ 16

Author(s):

Belén Rodríguez-Sánchez ◽

Emilia Cercenado ◽

Alix T. Coste ◽

Gilbert Greub

Keyword(s):

Public Health ◽

Control Measures ◽

Infection Prevention And Control ◽

Clinical Samples ◽

Hospital Hygiene ◽

Bacterial Typing ◽

Maldi Tof Ms ◽

Maldi Tof ◽

The Impact ◽

Tof Ms

Introduction MALDI-TOF MS represents a new technological era for microbiology laboratories. Improved sample processing and expanded databases have facilitated rapid and direct identification of microorganisms from some clinical samples. Automated analysis of protein spectra from different microbial populations is emerging as a potential tool for epidemiological studies and is expected to impact public health. Aim To demonstrate how implementation of MALDI-TOF MS has changed the way microorganisms are identified, how its applications keep increasing and its impact on public health and hospital hygiene. Methods A review of the available literature in PubMED, published between 2009 and 2018, was carried out. Results Of 9,709 articles retrieved, 108 were included in the review. They show that rapid identification of a growing number of microorganisms using MALDI-TOF MS has allowed for optimisation of patient management through prompt initiation of directed antimicrobial treatment. The diagnosis of Gram-negative bacteraemia directly from blood culture pellets has positively impacted antibiotic streamlining, length of hospital stay and costs per patient. The flexibility of MALDI-TOF MS has encouraged new forms of use, such as detecting antibiotic resistance mechanisms (e.g. carbapenemases), which provides valuable information in a reduced turnaround time. MALDI-TOF MS has also been successfully applied to bacterial typing. Conclusions MALDI-TOF MS is a powerful method for protein analysis. The increase in speed of pathogen detection enables improvement of antimicrobial therapy, infection prevention and control measures leading to positive impact on public health. For antibiotic susceptibility testing and bacterial typing, it represents a rapid alternative to time-consuming conventional techniques.

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