Instrument detects bacterial life forms

Instrument assays enzymatic bioluminescent reaction that occurs when adenosine triphosphate /ATP/ combines with lucifrase and luciferin. Module assembly minimizes need for hardware associated with reaction fluid and waste transfer. System is applicable in marine biology and aerospace and medical fields

Comparative analysis of pSMA198 found in Streptococcus macedonicus ACA-DC 198, the first streptococcal plasmid of the pCI305/pWV02 family of theta-replicating replicons

Here we analyze pSMA198, the first plasmid isolated from Streptococcus macedonicus ACA-DC 198, and we attempt to clarify the route of its original acquisition. Based on the similarity profiles of the plasmid’s replication initiation protein (Rep) and its origin of replication (ori), pSMA198 was found to be a novel member of the pCI305/pWV02 family of theta-replicating plasmids. The pCI305/pWV02 family consists of plasmids of narrow host range that are mainly found in lactococcal species. Comparative analysis of the pSMA198 revealed a high degree of similarity with plasmids pSK11b, pVF22 and pIL5 over its replication backbone, its mobilization backbone and most of its length, respectively. All these three plasmids have been isolated from Lactococcus lactis strains deriving from milk or its products supporting that S. macedonicus acquired pSMA198 from the latter species and that this acquisition took place in the dairy environment. Both pSMA198 and the chromosome of S. macedonicus exhibit a high degree of pseudogenes, indicating that they must have evolved under the same gene decay processes. Furthermore, we were able to determine chromosomal regions that may have originated from pSMA198, also supporting a long co-existence of the two replicons. In addition, pSMA198 is carried by S. macedonicus strains segregated in five different genotypes by pulsed-field gel electrophoresis (PFGE), showing that pSMA198’s acquisition is not a recent event. We propose that our overall analysis of pSMA198 points towards the habituation of S. macedonicus ACA-DC 198 to the dairy environment

Characterization of plasmid pSMA198 found in Streptococcus macedonicus ACA-DC 198 supports the relation of the species to the milk environment

Background: Streptococcus macedonicus is an intriguing streptococcal species whose most frequent source of isolation is fermented foods similarly to Streptococcus thermophilus. During the genome sequencing of S. macedonicus ACA-DC 198 a plasmid was identified. Objectives: To analyse pSMA198, the first plasmid isolated from S. macedonicus and to shed light onto its acquisition path. Methods: Similarity searches of nucleotide and protein sequences, comparative analysis of whole plasmid sequences and phylogenetic analysis were performed using the appropriate bioinformatics tools. Methods: Based on the similarity profiles of the plasmid’s replication initiation protein (Rep) and its origin of replication (ori), pSMA198 belongs to the narrow host range pCI305/pWV02 family found primarily in lactococci and it is the first such plasmid to be reported in streptococci. Comparative analysis of the pSMA198 over its ori, origin of transfer (oriT) or entire length revealed a high degree of similarity with plasmids pSK11b, pVF22 and pIL5, respectively, all isolated from Lactococcus lactis strains from milk or milk products. Phylogenetic analysis of the pSMA198 Rep showed that the vast majority of closely related proteins derive from lactococcal dairy isolates. Conclusions: Our findings demonstrate that S. macedonicus ACA-DC 198 acquired most probably plasmid pSMA198 from L. lactis during an ancestral genetic exchange event that took place in milk or dairy products. Based on our analysis we provide the first molecular and evolutionary evidence for the habituation of S. macedonicus to the dairy environment

Acquisition through Horizontal Gene Transfer of Plasmid pSMA198 by Streptococcus macedonicus ACA-DC 198 Points towards the Dairy Origin of the Species

Background: Streptococcus macedonicus is an intriguing streptococcal species whose most frequent source of isolation is fermented foods similarly to Streptococcus thermophilus. However, S. macedonicus is closely related to commensal opportunistic pathogens of the Streptococcus bovis/Streptococcus equinus complex. Methodology/Principal Findings: We analyzed the pSMA198 plasmid isolated from the dairy strain Streptococcus macedonicus ACA-DC 198 in order to provide novel clues about the main ecological niche of this bacterium. pSMA198 belongs to the narrow host range pCI305/pWV02 family found primarily in lactococci and to the best of our knowledge it is the first such plasmid to be reported in streptococci. Comparative analysis of the pSMA198 sequence revealed a high degree of similarity with plasmids isolated from Lactococcus lactis strains deriving from milk or its products. Phylogenetic analysis of the pSMA198 Rep showed that the vast majority of closely related proteins derive from lactococcal dairy isolates. Additionally, cloning of the pSMA198 ori in L. lactis revealed a 100% stability of replication over 100 generations. Both pSMA198 and the chromosome of S. macedonicus exhibit a high percentage of potential pseudogenes, indicating that they have co-evolved under the same gene decay processes. We identified chromosomal regions in S. macedonicus that may have originated from pSMA198, also supporting a long co-existence of the two replicons. pSMA198 was also found in divergent biotypes of S. macedonicus and in strains isolated from dispersed geographic locations (e.g. Greece and Switzerland) showing that pSMA198’s acquisition is not a recent event. Conclusions/Significance: Here we propose that S. macedonicus acquired plasmid pSMA198 from L. lactis via an ancestral genetic exchange event that took place most probably in milk or dairy products. We provide important evidence that point towards the dairy origin of this species

Multiplex imaging of live breast cancer tumour models through tissue using handheld surface enhanced spatially offset resonance Raman spectroscopy (SESORRS)

Through utilizing the depth penetration capabilities of SESORS, multiplexed imaging and classification of three singleplex nanotags and a triplex of nanotags within breast cancer tumour models is reported for the first time through depths of 10 mm using a handheld SORS instrument

Towards establishing a minimal nanoparticle concentration for applications involving surface enhanced spatially offset resonance Raman spectroscopy (SESORRS) in vivo

Resonant chalcogenpyrylium nanotags demonstrate an exceptional surface enhanced Raman scattering (SERS) performance for use in SORS applications. Using surface enhanced spatially offset Raman spectroscopy (SESORS), nanotags modified with a chalcogenpyrylium dye were observed at concentrations as low as 1 pM through 5 mm of tissue. Calculated limits of detection suggest that these SERS nanotags can be detected at 104 fM using surface enhanced spatially offset resonance Raman scattering (SESORRS) demonstrating their potential for in vivo applications

Surface enhanced resonance Raman spectroscopy (SERRS) for probing through plastic and tissue barriers using a handheld spectrometer

The ability to probe through barriers and tissue non-invasively is an urgent unmet need in both the security and biomedical imaging fields. Surface enhanced Raman spectroscopy (SERS) has been shown to yield superior enhancement in signal over conventional Raman techniques. Furthermore, by utilising a resonant Raman reporter to produce surface enhanced resonance Raman spectroscopy (SERRS), even greater enhancement in chemical signal can be generated. Here we show the benefit of using red-shifted chalcogenpyrylium based Raman reporters for probing through large thicknesses of plastic and tissue barriers using a conventional Raman instrument. Furthermore, the benefit of using a resonant Raman reporter for superior levels of through barrier detection is demonstrated, thus we aim to show the advantage of using resonant nanotags in combination with conventional Raman spectroscopy to probe through plastic and tissue barriers. Raman signals were collected from SERRS active nanotags through plastic thicknesses of up to 20 mm, as well as the detection of the same SERRS nanotags through up to 10 mm of tissue sections using a handheld conventional Raman spectrometer. The ability to detect SERRS-active nanotags taken up into ex vivo tumour models known as multicellular tumour spheroids (MTS), through depths of 5 mm of tissue was also shown. The advantages of applying multivariate analysis for through barrier detection when discriminating analytes with similar spectral features as the barrier is also clearly demonstrated. To the best of our knowledge, this is the first report of the assessment of the maximum level of through barrier detection using a conventional handheld Raman instrument for SERS applications as well as demonstration of the power of resonant nanotags for probing through barriers using conventional Raman spectroscopy

Cellular and Transcriptional Responses of Crassostrea gigas Hemocytes Exposed in Vitro to Brevetoxin (PbTx-2)

Hemocytes mediate a series of immune reactions essential for bivalve survival in the environment, however, the impact of harmful algal species and their associated phycotoxins upon bivalve immune system is under debate. To better understand the possible toxic effects of these toxins, Crassostrea gigas hemocytes were exposed to brevetoxin (PbTx-2). Hemocyte viability, monitored through the neutral red retention and MTT reduction assays, and apoptosis (Hoechst staining) remained unchanged during 12 h of exposure to PbTx-2 in concentrations up to 1000 µg/L. Despite cell viability and apoptosis remained stable, hemocytes incubated for 4 h with 1000 µg/L of PbTx-2 revealed higher expression levels of Hsp70 (p < 0.01) and CYP356A1 (p < 0.05) transcripts and a tendency to increase FABP expression, as evaluated by Real-Time quantitative PCR. The expression of other studied genes (BPI, IL-17, GSTO, EcSOD, Prx6, SOD and GPx) remained unchanged. The results suggest that the absence of cytotoxic effects of PbTx-2 in Crassostrea gigas hemocytes, even at high concentrations, allow early defense responses to be produced by activating protective mechanisms associated to detoxification (CYP356A1 and possibly FABP) and stress (Hsp70), but not to immune or to antioxidant (BPI, IL-17, EcSOD, Prx6, GPx and SOD) related genes

Concurrent Exposure of Bottlenose Dolphins (Tursiops truncatus) to Multiple Algal Toxins in Sarasota Bay, Florida, USA

Sentinel species such as bottlenose dolphins (Tursiops truncatus) can be impacted by large-scale mortality events due to exposure to marine algal toxins. In the Sarasota Bay region (Gulf of Mexico, Florida, USA), the bottlenose dolphin population is frequently exposed to harmful algal blooms (HABs) of Karenia brevis and the neurotoxic brevetoxins (PbTx; BTX) produced by this dinoflagellate. Live dolphins sampled during capture-release health assessments performed in this region tested positive for two HAB toxins; brevetoxin and domoic acid (DA). Over a ten-year study period (2000–2009) we have determined that bottlenose dolphins are exposed to brevetoxin and/or DA on a nearly annual basis (i.e., DA: 2004, 2005, 2006, 2008, 2009; brevetoxin: 2000, 2004, 2005, 2008, 2009) with 36% of all animals testing positive for brevetoxin (n = 118) and 53% positive for DA (n = 83) with several individuals (14%) testing positive for both neurotoxins in at least one tissue/fluid. To date there have been no previously published reports of DA in southwestern Florida marine mammals, however the May 2008 health assessment coincided with a Pseudo-nitzschia pseudodelicatissima bloom that was the likely source of DA observed in seawater and live dolphin samples. Concurrently, both DA and brevetoxin were observed in common prey fish. Although no Pseudo-nitzschia bloom was identified the following year, DA was identified in seawater, fish, sediment, snails, and dolphins. DA concentrations in feces were positively correlated with hematologic parameters including an increase in total white blood cell (p = 0.001) and eosinophil (p<0.001) counts. Our findings demonstrate that dolphins within Sarasota Bay are commonly exposed to two algal toxins, and provide the impetus to further explore the potential long-term impacts on bottlenose dolphin health