Material and Methods: This experimental study involved 225 bone specimens prepared from discarded bone fragments during a series of 45 knee and hip arthroplasty surgeries. The bone fragments were cut into five identical cubes and were randomly assigned to either control (positive or negative), or experimental groups (0.5% chlorhexidine, 10% povidone-iodine or 70% alcohol). The control negative was to determine pre-contamination culture. All bone specimens, except the control negative group were uniformly contaminated by dropping on the operation theatre floor. Subsequently, the dropped bone specimens except for the control positive group, were disinfected by immersing in a respective antiseptic solution for 10 minutes, before transported to the microbiology laboratory for incubation.
Results: The incidence of a positive culture from a dropped bone fragment was 86.5%. From the 37 specimens sent for each group, the incidence of positive culture was 5.4% (2 specimens) after being disinfected using chlorhexidine, 67.6% (25 specimens) using povidone-iodine and 81.1% (30 specimens) using alcohol. Simple logistic regression analysis demonstrated that chlorhexidine was significantly effective in disinfecting contaminated bones (p-value <0.001, odd ratio 0.009). Povidone-iodine and alcohol were not statistically significant (p-value 0.059 and 0.53, respectively). Organisms identified were Bacillus species and coagulase negative Staphylococcus. No gram-negative bacteria were isolated.
Conclusion: A total of 0.5% chlorhexidine is effective and superior in disinfecting contaminated bones.
METHODS: The enzyme was purified in two steps using affinity and size exclusion chromatography. Enzyme assays were performed using the malachite green assay and enzymatic product was identified using gas chromatography-mass spectrometry (GC-MS) analysis. Sequence analysis of PmSTS was performed using multiple sequence alignment (MSA) against plant sesquiterpene synthase sequences. The homology model of PmSTS was generated using I-TASSER server.
RESULTS: Our findings suggest that the recombinant PmSTS is mainly expressed as inclusion bodies and soluble aggregate in the E. coli protein expression system. However, the addition of 15% (v/v) glycerol to the protein purification buffer and the removal of N-terminal 24 amino acids of PmSTS helped to produce homogenous recombinant protein. Enzyme assay showed that recombinant PmSTS is active and specific to the C15 substrate FPP. The optimal temperature and pH for the recombinant PmSTS are 30 °C and pH 8.0, respectively. The GC-MS analysis further showed that PmSTS produces β-sesquiphellandrene as a major product and β-farnesene as a minor product. MSA analysis revealed that PmSTS adopts a modified conserved metal binding motif (NSE/DTE motif). Structural analysis suggests that PmSTS may binds to its substrate similarly to other plant sesquiterpene synthases.
DISCUSSION: The study has revealed that homogenous PmSTS protein can be obtained with the addition of glycerol in the protein buffer. The N-terminal truncation dramatically improved the homogeneity of PmSTS during protein purification, suggesting that the disordered N-terminal region may have caused the formation of soluble aggregate. We further show that the removal of the N-terminus disordered region of PmSTS does not affect the product specificity. The optimal temperature, optimal pH, Km and kcat values of PmSTS suggests that PmSTS shares similar enzyme characteristics with other plant sesquiterpene synthases. The discovery of an altered conserved metal binding motif in PmSTS through MSA analysis shows that the NSE/DTE motif commonly found in terpene synthases is able to accommodate certain level of plasticity to accept variant amino acids. Finally, the homology structure of PmSTS that allows good fitting of substrate analog into the catalytic active site suggests that PmSTS may adopt a sesquiterpene biosynthesis mechanism similar to other plant sesquiterpene synthases.
METHODS: In this study, we extracted retrospective data from Malaysian surveillance database. Analysis was carried out using WHONET software focusing noncholera Vibrio spp. including Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio fluvialis, Vibrio alginolyticus, Vibrio hollisae (Grimontia hollisae), Vibrio mimicus, Vibrio metschnikovii, and Vibrio furnissii.
RESULTS: Here, we report the first distribution and prevalence of these species isolated in Malaysia together with the antibiotic sensitivity profile based on the species. We found that V. parahaemolyticus is the predominant species isolated in Malaysia. Noticeably, across the study period, V. fluvialis is becoming more prevalent, as compared to V. parahaemolyticus. In addition, this study also reports the first isolation of pathogenic V. furnissii from stool in Malaysia.
CONCLUSION: These data represent an important step toward understanding the potential emergence of noncholera Vibrio spp. outbreaks.
CASE REPORT: A 3-month-old baby boy was presented with neonatal anaemia and mild hepatomegaly. Full blood count revealed severe hypochromic microcytic anaemia. There was an abundance of HbH inclusion bodies in his red blood cells. High-performance liquid chromatography showed a reduced HbA2 level with the presence of pre-run peak. Capillary electrophoresis showed the presence of HbH and Hb Barts. Molecular analysis found a common α0-thalassaemia (--SEA) in one allele and mutation in codon 125 in the other allele.
DISCUSSION: Non-deletional HbH disease due to a combination of deletional and non-deletional mutations may present with severe clinical manifestations than those with deletion mutations, which warrants accurate diagnosis using molecular techniques.