Intracellular polyphosphate length characterization in polyphosphate accumulating microorganisms (PAOs): Implications in PAO phenotypic diversity and enhanced biological phosphorus removal performance

Polyphosphate (polyP) accumulating organisms (PAOs) are the key agent to perform enhanced biological phosphorus removal (EBPR) activity, and intracellular polyP plays a key role in this process. Potential associations between EBPR performance and the polyP structure have been suggested, but are yet to be extensively investigated, mainly due to the lack of established methods for polyP characterization in the EBPR system. In this study, we explored and demonstrated that single-cell Raman spectroscopy (SCRS) can be employed for characterizing intracellular polyPs of PAOs in complex environmental samples such as EBPR systems. The results, for the first time, revealed distinct distribution patterns of polyP length (as Raman peak position) in PAOs in lab-scale EBPR reactors that were dominated with different PAO types, as well as among different full-scale EBPR systems with varying configurations. Furthermore, SCRS revealed distinctive polyP composition/features among PAO phenotypic sub-groups, which are likely associated with phylogenetic and/or phenotypic diversity in EBPR communities, highlighting the possible resolving power of SCRS at the microdiversity level. To validate the observed polyP length variations via SCRS, we also performed and compared bulk polyP length characteristics in EBPR biomass using conventional polyacrylamide gel electrophoresis (PAGE) and solution 31P nuclear magnetic resonance (31P-NMR) methods. The results are consistent with the SCRS findings and confirmed the variations in the polyP lengths among different EBPR systems. Compared to conventional methods, SCRS exhibited advantages as compared to conventional methods, including the ability to characterize in situ the intracellular polyPs at subcellular resolution in a label-free and non-destructive way, and the capability to capture subtle and detailed biochemical fingerprints of cells for phenotypic classification. SCRS also has recognized limitations in comparison with 31P-NMR and PAGE, such as the inability to quantitatively detect the average polyP chain length and its distribution. The results provided initial evidence for the potential of SCRS-enabled polyP characterization as an alternative and complementary microbial community phenotyping method to facilitate the phenotype-function (performance) relationship deduction in EBPR systems

The quantitative assessment of UHMWPE wear debris produced in hip simulator testing : the influence of head material and roughness, motion and loading.

Biomaterial wear particles are known to provoke a foreign-body reaction when released from total joint prostheses. Considerable effort is being invested in the search for materials which, by wearing less, will release fewer particles. In this research the role of joint wear simulators is paramount. The wear debris from 26 simulator tests of hip prostheses was extracted from the bovine serum lubricant and sized using a laser diffraction technique. The influence on particle size of a broad range of parameters was examined. The parameters considered included the bedding-in phenomenon, the femoral head material and roughness and physiological versus simplified load cycles and wear paths. The physiological wear simulators produced a similar size distribution of wear particles to that produced by implanted joints. Head material had no effect on this observation. Simplifying the loading cycle or wear path independently made no impact on this finding. This remained the case when simplified load and wear path were combined in one test. The effect of head roughness was pronounced with an increase in minimum particle size with increasing roughness. Joint simulators remain the optimal method for assessing new joint materials and designs. However, their use to characterise joints in terms of wear rate solely should be guarded against. Instead focus should be concentrated on a combination of the size and amount of the wear debris created. In this way the "loosening hazard" of a joint can be distilled