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Characterization of bacteria in ballast water using MALDI-TOF mass spectrometry.

Emami K, Askari V, Ullrich M, Mohinudeen K, Anil AC, Khandeparker L, Burgess JG, Mesbahi E - PLoS ONE (2012)

Bottom Line: To evaluate a rapid and cost-effective method for monitoring bacteria in ballast water, several marine bacterial isolates were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).Seawater samples collected from the North Sea were incubated in steel ballast tanks and the presence of potentially harmful species of Pseudomonas was also investigated.At the genus-level, the identification of thirty six isolates using MALDI-TOF MS produced similar results to those obtained by 16S rRNA gene sequencing.

View Article: PubMed Central - PubMed

Affiliation: School of Biology, Newcastle University, Newcastle upon Tyne, United Kingdom. kaveh.emami@ncl.ac.uk

ABSTRACT
To evaluate a rapid and cost-effective method for monitoring bacteria in ballast water, several marine bacterial isolates were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Since International Maritime Organization (IMO) regulations are concerned with the unintended transportation of pathogenic bacteria through ballast water, emphasis was placed on detecting species of Vibrio, enterococci and coliforms. Seawater samples collected from the North Sea were incubated in steel ballast tanks and the presence of potentially harmful species of Pseudomonas was also investigated. At the genus-level, the identification of thirty six isolates using MALDI-TOF MS produced similar results to those obtained by 16S rRNA gene sequencing. No pathogenic species were detected either by 16S rRNA gene analysis or by MALDI-TOF MS except for the opportunistically pathogenic bacterium Pseudomonas aeruginosa. In addition, in house software that calculated the correlation coefficient values (CCV) of the mass spectral raw data and their variation was developed and used to allow the rapid and efficient identification of marine bacteria in ballast water for the first time.

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MALDI-TOF mass spectrograms, (panel A) and 16S rRNA gene sequence phylogram, (panel B) for four Vibrio isolates from the North Sea.Panel A, the X axis indicates the mass to charge ratios (m/z) of each peak and the Y axis for each individual plot indicates intensity of the peaks. The identification result using Biotyper software is shown at the top-right corner of each plot followed by the type strain designation. Several mass spectral peaks are common between the isolates and a few show differences, examples of similar peaks are aligned by dotted lines. Panel B, distance phylogram tree of the isolates, over 1500 nucleotides.
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pone-0038515-g001: MALDI-TOF mass spectrograms, (panel A) and 16S rRNA gene sequence phylogram, (panel B) for four Vibrio isolates from the North Sea.Panel A, the X axis indicates the mass to charge ratios (m/z) of each peak and the Y axis for each individual plot indicates intensity of the peaks. The identification result using Biotyper software is shown at the top-right corner of each plot followed by the type strain designation. Several mass spectral peaks are common between the isolates and a few show differences, examples of similar peaks are aligned by dotted lines. Panel B, distance phylogram tree of the isolates, over 1500 nucleotides.

Mentions: Vibrio species are commonly found in seawater and are of special interest due to their potential to cause specific diseases, such as gastroenteritis, cholera, sepsis, and cellulitis leading to diarrhea and necrotizing soft tissue infections [26], [27]. Due to increased global marine traffic, the spread of potentially harmful Vibrio species deserves particular attention on shores of highly populated European countries [28]. IMO is also specifically concerned about the concentrations of serotypes O1 and O139 of V. cholerae in ballast water [6]. V. cholerae was not detected in this study. Four out of eight Vibrio isolates in this study, show high similarities at the nucleotide and protein levels. MTB analysis of the isolates S32, S27, S14 and S30 by FlexAnalysis software revealed that they all contain major peaks with similar m/z and relative intensities. MS data of isolates S14, S32, and S27 followed by Biotyper software analysis supported the possibility that isolates S14 and S32 belonged to the same species, V. gigantis (Fig. 1). By comparing the 16S rRNA gene sequences of all four Vibrio isolates using the ClustalW2 alignment program, it was found that the differences were not significant enough to confidently differentiate them into separate species (Table 2). 16S rRNA gene comparison and BLAST search however implied that isolates S27 and S30 are similar to V. cyclitrophicus. Isolate S30 was also identified as V. cyclitrophicus by MALDI-TOF analysis and Biotyper software. However, the identity of the nucleotide sequence of S30 to that of the type strain of V. cyclitrophicus in NCBI database was only 98%. Likewise, relatively low Biotyper score of 2.1 out of 3.0 indicated uncertainty in the species detection by both methods. There were few differences in mass-spectra of the four above mentioned isolates, for example, the mass spectrum of isolate S27 contained peaks at m/z 8010 and 6653, which were missing in isolates S14 and S32. Peaks at m/z 3973 and 3976 in isolates S32 and S14 were missing in isolate S27 (Fig. 1). Although some of the differences between closely related species may be due to amino acid substitutions, the m/z differences between these isolates did not indicate any altered amino acid sequences.


Characterization of bacteria in ballast water using MALDI-TOF mass spectrometry.

Emami K, Askari V, Ullrich M, Mohinudeen K, Anil AC, Khandeparker L, Burgess JG, Mesbahi E - PLoS ONE (2012)

MALDI-TOF mass spectrograms, (panel A) and 16S rRNA gene sequence phylogram, (panel B) for four Vibrio isolates from the North Sea.Panel A, the X axis indicates the mass to charge ratios (m/z) of each peak and the Y axis for each individual plot indicates intensity of the peaks. The identification result using Biotyper software is shown at the top-right corner of each plot followed by the type strain designation. Several mass spectral peaks are common between the isolates and a few show differences, examples of similar peaks are aligned by dotted lines. Panel B, distance phylogram tree of the isolates, over 1500 nucleotides.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3369924&req=5

pone-0038515-g001: MALDI-TOF mass spectrograms, (panel A) and 16S rRNA gene sequence phylogram, (panel B) for four Vibrio isolates from the North Sea.Panel A, the X axis indicates the mass to charge ratios (m/z) of each peak and the Y axis for each individual plot indicates intensity of the peaks. The identification result using Biotyper software is shown at the top-right corner of each plot followed by the type strain designation. Several mass spectral peaks are common between the isolates and a few show differences, examples of similar peaks are aligned by dotted lines. Panel B, distance phylogram tree of the isolates, over 1500 nucleotides.
Mentions: Vibrio species are commonly found in seawater and are of special interest due to their potential to cause specific diseases, such as gastroenteritis, cholera, sepsis, and cellulitis leading to diarrhea and necrotizing soft tissue infections [26], [27]. Due to increased global marine traffic, the spread of potentially harmful Vibrio species deserves particular attention on shores of highly populated European countries [28]. IMO is also specifically concerned about the concentrations of serotypes O1 and O139 of V. cholerae in ballast water [6]. V. cholerae was not detected in this study. Four out of eight Vibrio isolates in this study, show high similarities at the nucleotide and protein levels. MTB analysis of the isolates S32, S27, S14 and S30 by FlexAnalysis software revealed that they all contain major peaks with similar m/z and relative intensities. MS data of isolates S14, S32, and S27 followed by Biotyper software analysis supported the possibility that isolates S14 and S32 belonged to the same species, V. gigantis (Fig. 1). By comparing the 16S rRNA gene sequences of all four Vibrio isolates using the ClustalW2 alignment program, it was found that the differences were not significant enough to confidently differentiate them into separate species (Table 2). 16S rRNA gene comparison and BLAST search however implied that isolates S27 and S30 are similar to V. cyclitrophicus. Isolate S30 was also identified as V. cyclitrophicus by MALDI-TOF analysis and Biotyper software. However, the identity of the nucleotide sequence of S30 to that of the type strain of V. cyclitrophicus in NCBI database was only 98%. Likewise, relatively low Biotyper score of 2.1 out of 3.0 indicated uncertainty in the species detection by both methods. There were few differences in mass-spectra of the four above mentioned isolates, for example, the mass spectrum of isolate S27 contained peaks at m/z 8010 and 6653, which were missing in isolates S14 and S32. Peaks at m/z 3973 and 3976 in isolates S32 and S14 were missing in isolate S27 (Fig. 1). Although some of the differences between closely related species may be due to amino acid substitutions, the m/z differences between these isolates did not indicate any altered amino acid sequences.

Bottom Line: To evaluate a rapid and cost-effective method for monitoring bacteria in ballast water, several marine bacterial isolates were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).Seawater samples collected from the North Sea were incubated in steel ballast tanks and the presence of potentially harmful species of Pseudomonas was also investigated.At the genus-level, the identification of thirty six isolates using MALDI-TOF MS produced similar results to those obtained by 16S rRNA gene sequencing.

View Article: PubMed Central - PubMed

Affiliation: School of Biology, Newcastle University, Newcastle upon Tyne, United Kingdom. kaveh.emami@ncl.ac.uk

ABSTRACT
To evaluate a rapid and cost-effective method for monitoring bacteria in ballast water, several marine bacterial isolates were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Since International Maritime Organization (IMO) regulations are concerned with the unintended transportation of pathogenic bacteria through ballast water, emphasis was placed on detecting species of Vibrio, enterococci and coliforms. Seawater samples collected from the North Sea were incubated in steel ballast tanks and the presence of potentially harmful species of Pseudomonas was also investigated. At the genus-level, the identification of thirty six isolates using MALDI-TOF MS produced similar results to those obtained by 16S rRNA gene sequencing. No pathogenic species were detected either by 16S rRNA gene analysis or by MALDI-TOF MS except for the opportunistically pathogenic bacterium Pseudomonas aeruginosa. In addition, in house software that calculated the correlation coefficient values (CCV) of the mass spectral raw data and their variation was developed and used to allow the rapid and efficient identification of marine bacteria in ballast water for the first time.

Show MeSH
Related in: MedlinePlus