IN-FOCUS: Gumming up the works: oral biofilms under the microscope.

Image showing yeast cells colonising a tissue-engineered oral epithelium

Yeast cells (Candida albicans; red) colonise and invade a tissue-engineered oral epithelium (cell nuclei; blue; cytoplasm, green)

The BIOSI Bioimaging Facility has worked closely with Professor David Williams at the Dental School in Cardiff for a number of years. David is an expert in oral microbiology, specialising in microbial biofilms (e.g. dental plaque) and mycoses such as oral candidiases (thrush). Over this time, we have been involved in a number of collaborative studies where we have used confocal microscopy and various fluorescent labelling techniques to investigate the formation, 3D organization and microbial community structure of biofilms grown on tissue engineered oral epithelium, endotracheal tubes and substrates such as dental acrylic and titanium. The research has also evaluated the effect of various anti-microbial and anti-fungal compounds and commercial mouth rinses on biofilm development using fluorescent viability stains. The studies have extended our understanding of how oral biofilms develop and in how they respond to therapeutic intervention, and have resulted in a number of publications (see below) as well as a book cover for a leading text on the subject of Oral Microbiology. It’s a fantastic application of confocal microscopy to a biological problem and, from an imaging perspective, its been something for us to really get our teeth into!

AJH

Find out more:
Further reading:

IN-FOCUS: Shedding light on new fluors: development of novel, metal-based probes for bioimaging.

Dr Simon Pope and his research group in CHEMY have been collaborating with the BIOSI Bioimaging Unit for over 8 years. Their research is focussed upon the development of new, metal-based, fluorescent probes for cell imaging applications, and forms part of a larger study on the use of metal complexes, which include rhenium and gold, as multi-modal imaging agents with therapeutic potential. In these studies, we have performed confocal imaging to (1) assess the cytotoxicity of the new probes, (2) evaluate their cellular uptake and determine their cytoplasmic localisation, and (3) characterise their fluorescent emissions via spectral (wavelength or lambda) scanning. The collaboration has yielded a number of high impact publications (see below) as well as a journal cover! With the improved potential of the new super-resolution confocal system we anticipate a lot more to come.

AJH

Find out more:

NEWS: Naomi’s nightmares of nature.

Naomi

Image: BBC camera crew (left) filming the follicle mite, Demodex (right), for an episode of the children’s television programme, Naomi’s Nightmare’s of Nature.

Not so long ago we received a strange request from Dr Sarah Perkins (BIOSI): could we accommodate a BBC camera crew within the Bioimaging Facility to film an episode of the Children’s CBBC television programme, Naomi’s Nightmares of Nature? The nightmare in question, was the eyelash mite,  Demodex, a commensal ectoparasite that lives within the hair follicles (Demodex follicularum) and sebaceous glands (Demodex brevis) of the face, feeding off sebum and other organic detritus. Anyway, prior to filming, we spent an anxious morning attempting to isolate live Demodex from ‘volunteers’ faces by various means,  including via sellotape, with little success Fortunately, when Naomi and her production team arrived, we struck gold! A few eyelashes extracted from their cameraman, Steve, revealed a bumper load of parasites and, using DIC optics, we were able to generate some nice microscopic footage of a family of mites tucking into their evening meal!

AJH

Find out more:

IN-FOCUS: Heavy metal worms under the spotlight.

chloragocyte

Not just pretty pictures: an earthworm coelomocyte imaged by conventional confocal microscopy (left; DIC/fluorescence image) and via spectral scanning (centre) to analyse changes in riboflavin fluorescence caused by soil pollution (right).

The Research Techniques module run by Professor Pete Kille and Dr Carsten Muller (Introduction to Environmental Toxicology) makes for a busy week within the Bioimaging Research Hub. In the practical, students learn a range of advanced analytical research techniques as they aim to identify and characterize earthworm populations that have been sampled from land polluted by heavy metal – and I’m not talking about Axel’s Rose garden here : )

Pete is an expert in ecotoxicology and much of his research centers on how invertebrate species, such as the earthworm, deal with heavy metal pollutants, e.g., lead, in their environment. As it turns out, they seem to be pretty good at tolerating a lot of the nasty stuff that passes through them, but it does leave an indelible metabolic mark – making the organisms ideal for environmental toxicological testing. And here’s where it gets interesting: previous studies of the earthworm, Eisenia fetida, have shown that heavy metals affect riboflavin (vitamin B2) biosynthesis. Now, riboflavin happens to be (1) highly autofluorescent, and (2) neatly packaged within spheroidal organelles, or chloragosomes, within a sub-population of  immune cells, called coelomocytes, that are resident within the body cavity of the worm. Fortunately, earthworms can be gently persuaded to give up some of these cells for confocal microscopic analysis.

In the practical, we use confocal microscopy to image earthworm coelomocytes and, via spectral scanning, generate emission spectra of the riboflavin autofluorescence from within the chloragosomes. By comparing the autofluorescent signatures of coelomocytes from worms obtained from different sampling sites, we have asked the question: can riboflavin autofluorescence in this organism be used to assess soil pollution?

And the answer? Well, I’m not at liberty to say – the students reports aren’t in yet! (Answers on the back of a postcard to…)

AJH

Find out more:

Zimmerman et al. (2003) Spectral Imaging and its applications in live cell microscopy. FEBS Letters 546: 87-92.

Further Reading

SPOKE EQUIPMENT: New Multi-Spectral In Vivo Imaging system.

Dr Amit Jathoul and Professor Jim Murray (BIOSI) have recently obtained generous funding from the Wellcome Trust to purchase a real-time, multi-spectral in vivo imaging system from Biospace Lab. The PhotonIMAGER allows bioluminescence/fluorescence imaging at a wide range of scales from cells, tissues and organs to entire complex organisms, thereby bridging the gap  from single cell to whole organism imaging. Further details of the system are available through the research equipment database.

AJH

Find out more:
Further reading:

IN-FOCUS: Imaging the Mitten crab: working hand in glove with the Natural History Museum

Imaris Snapshot

3D surface-rendered model of appendage (endopod of maxilliped) of Chinese mitten crab larva. Image produced on Zeiss LSM880 confocal and reconstructed using Bitplane Imaris.

Last week we undertook some confocal microscopy for the National History Museum to help characterise the arrangement of setae on the larval appendages of the Chinese mitten crab, Eriocheir sinesis * (now published, see Kamanli et al, 2017 below). The Mitten crab, so-named because of the tufts of ‘fur’ on the adult’s claws, is officially listed as one of the World’s most invasive species. The crabs out-compete and prey on native crab species, damage fishing nets and cause significant erosion of riverbanks, thus are of considerable economic importance.  They arrived in this country from China in the 1930’s via discharge of ballast water from trading ships and are now firmly established in many of Britain’s waterways. The National History Museum is investigating ways of reducing the population of Mitten crabs and this species is currently under evaluation as a potential food source in the UK (so if you can’t beat them, eat them!)

AJH

Find out more:
Further reading:

 

IN FOCUS: Development of a Virtual Histology Slide Box.

histology database & viewer

Image: The virtual histology slide box and viewer – a resource that holds fantastic potential for BIOSI teaching, research and public engagement.

The Bioimaging Hub has recently completed work in digitising the School’s extensive histopathology slide repository. Over 400 histological sections, encompassing both normal and pathological tissues, were painstakingly scanned and digitised in high resolution using the facilities Objective Imaging Surveyor slide scanning system.  The datasets, totalling 4TB, have been converted into the Zoomify .ziff image file format to enable easy and rapid on-line browsing, zooming and navigation (similar to that of Google Earth) and calibrated to allow feature measurement. The image files have been linked, via thumbnails, to a database that captures all relevant metadata for each histological section (filename, tissue type, organ system, species, section plane, histological stain, section ID, supplier, objective magnification etc) to facilitate easy sorting and data retrieval. The database is currently set up on a basic Linux server within the facility; however, to cope with concurrent file access by large numbers of up to 150 students, it will require a permanent home  on a dedicated server within the School. With further development, the resource promises to have fantastic potential for teaching, research and public engagement within BIOSI. Thanks to all concerned who have taken the project this far…

AJH

Find out more:

 

CORE EQUIPMENT: New Bitplane Imaris Workstation for Advanced 3D/4D Image Processing.

Imaris Screenshot

Image: Screenshot of the new Imaris software showing a surface-rendered model of an insect’s compound eye.

The Bioimaging Research Hub has recently purchased Bitplane Imaris software for Cell Biologists, together with a high-end PC workstation, via generous funding from the Research Infrastructure Fund (lead applicant: Dr Walter Dewitte). The software provides advanced  processing options for confocal and multi-photon 3D/4D datasets and includes the following modules: Measurement Pro, Imaris Track, Imaris Cell, Imaris Coloc, Imaris XT and Imaris Vantage. Further details of the system are available via our equipment database.

AJH

Find out more:

EQUIPMENT: New State-of-the-Art Confocal Microscope with Super-Resolution Capability.

DSC_0459

Image: The old and the new: The new Zeiss LSM 880 Airyscan confocal system (right) adjacent to the old Leica TCS SP2 confocal (left) (BIOSI 2; E/0.03)

The BIOSI Bioimaging Research Hub has recently expanded its imaging toolbox with a new, state-of-the-art confocal microscope system, that was purchased through generous funding by the Research Infrastructure Fund (Lead Applicant: Dr Walter Dewitte). The system, a top-of-the-range Zeiss LSM 880 upright confocal microscope, features the advanced Airyscan super-resolution detection module which provides a 1.7x gain in resolution in all three dimensions compared to conventional confocal optics. The system also supports advanced fluorescence techniques including FCS (fluorescence correlation microscopy) and FLIM (fluorescence lifetime imaging (FLIM) – the FLIM module will be installed during the first week of December. Further details here.

AJH

Find out more: