T. K. Chan

A prospective study of symptomatic bacteremia following platelet transfusion and of its management

BACKGROUND: The danger of bacteremia due to contaminated platelets is not well known. There are also no established guidelines for the management of febrile reactions after platelet transfusion.

Treatment of adenovirus-associated haemorrhagic cystitis with ganciclovir

We report a 47-year-old bone marrow transplant recipient with haemorrhagic cystitis caused by adenovirus successfully treated with ganciclovir. This is the first report on the use of ganciclovir for the successful treatment of adenoviral infection. Shell vial culture may be more sensitive than conventional culture in the detection of adenovirus in such patients.

Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Rapid Identification of Burkholderia pseudomallei: Importance of Expanding Databases with Pathogens Endemic to Different Localities

Burkholderia pseudomallei is the causative agent of melioidosis, a serious disease endemic in southeast Asia. Accurate identification of B. pseudomallei is important, since treatment of melioidosis requires prolonged antibiotics to prevent relapse ([9][1]). Although B. pseudomallei differs greatly

Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry for identification of clinically significant bacteria that are difficult to identify in clinical laboratories

Aims Although the revolutionary matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) has been evaluated for identification of various groups of bacteria, its application in bacteria that are ‘difficult-to-identify’ by phenotypic tests has been less well studied. We aim to evaluate the usefulness of MALDI-TOF MS for identification of ‘difficult-to-identify’ bacterial isolates. Methods We evaluated the performance of the Bruker MALDI-TOF MS system for a collection of 67 diverse clinically important bacterial isolates that were less commonly encountered, possessed ambiguous biochemical profiles or belonged to newly discovered species. The results were compared with 16S rRNA gene sequencing as a reference method for species identification. Results Using 16S rRNA gene sequencing as the reference method, 30 (45%) isolates were identified correctly to species level (score ≥2.0), 20 (30%) were only identified to genus level (score ≥1.7), four (6%) were misidentified (incorrect species with score ≥2.0 or incorrect genus with score ≥1.7) and 13 (19%) showed ‘no identification’ (score <1.7). Aerobic Gram-positive bacteria showed the highest percentage of correct species identification, followed by aerobic Gram-negative, anaerobic Gram-positive and anaerobic Gram-negative bacteria. Sixteen isolates identified to genus level actually showed the correct species but with scores below the threshold for species identification. Most isolates which showed ‘no identification’ were due to the absence of the corresponding species in the Bruker database. Conclusions Expansion of commercial databases to include reference spectra of less commonly encountered and newly discovered species and to increase available spectra for each species is required to improve the accuracy of MALDI-TOF MS for identifying ‘difficult-to-identify’ bacteria.

Pulmonary infections in bone marrow transplantation : the Hong Kong experience

The pattern of pulmonary infections in 59 consecutive bone marrow transplantations in Hong Kong was reviewed. Compared with published data from other marrow transplant units, we had a lower incidence of cytomegalovirus pneumonitis (1.7%) and a higher incidence of mycobacterial infections (5%). The latter is probably related to the high background prevalence of tuberculosis in the local population. Treatment with antituberculous drugs was effective.

Detection of CMV DNA in bone marrow transplant recipients: plasma versus leucocyte polymerase chain reaction.

AIMS: To compare the nested polymerase chain reaction (PCR) assay for the detection of cytomegalovirus (CMV) DNA in peripheral blood leucocytes and plasma obtained from heparinised blood; to determine the efficiency of various DNA extraction methods to minimise inhibition of plasma PCR and their effect on the sensitivity of plasma PCR; to determine the inhibitory effect of heparin, dextran, and EDTA on the CMV PCR assay. METHODS: 217 heparinised blood specimens from 58 bone marrow transplant patients were processed and the sensitivities and specificities of the PCR assays using peripheral blood leucocytes and plasma (with simple, Instagene, and Geneclean extraction methods) were compared to those of conventional CMV culture. In a separate experiment, dilutions of heparin, dextran, and EDTA were included in PCR assays. RESULTS: The detection of CMV DNA using peripheral blood leucocytes for PCR assay was significantly more sensitive (100%) than when using plasma (60%). Instagene and Geneclean extraction removed inherent inhibition but did not improve the sensitivity of the plasma PCR reaction. Heparin had an inhibitory effect on PCR. CONCLUSIONS: PCR assay using peripheral blood leucocytes is better than plasma for guiding the prescription of ganciclovir to bone marrow transplant patients. Heparin is inhibitory to the plasma PCR reaction.

Complementary use of MALDI-TOF MS and real-time PCR–melt curve analysis for rapid identification of methicillin-resistant staphylococci and VRE

OBJECTIVESnTo develop a rapid method for routine screening of methicillin-resistant staphylococci and VRE for clinical isolates and positive blood cultures.nnnMETHODSnOur method consisted of two parts: MALDI-TOF MS was used for identification of staphylococci and enterococci, followed by antibiotic resistance detection by real-time PCR-melt curve analysis without DNA extraction. The latter part included a triplex reaction for staphylococcal culture isolates (mecA, mecALGA251 and Panton-Valentine leucocidin genes), dual PCR of mecA/mecALGA251 and nuc genes for staphylococcal blood cultures, and a duplex reaction for enterococci (vanA and vanB genes). A total of 124 clinical isolates and 56 positive blood cultures were tested. MALDI-TOF MS was performed using Microflex LT (Bruker Daltonik, Bremen, Germany) and Rotor-Gene Q (Qiagen, Hilden, Germany) was used for real-time PCR-melt curve analysis. The total assay time was <2.5 h.nnnRESULTSnThe results revealed 100% concordance with antibiotic susceptibility testing or other reference methods for all culture isolates and enterococcal blood cultures. The percentage of concordance for staphylococcal blood cultures was 97.5%.nnnCONCLUSIONSnThe method described herein was fast, economical, reliable and capable of detecting mecALGA251, vanB1 and vanB2 genotypes, which are not included in most commercial assays. Large-scale screening is required to further test the performance of this protocol, especially for genotypes that are infrequently encountered.

Matrix-assisted laser desorption ionisation–time of flight mass spectrometry for rapid identification of Laribacter hongkongensis

Laribacter hongkongensis is a Gram-negative, facultative anaerobic, motile, S-shaped, urease-positive bacillus associated with invasive infections in liver cirrhosis patients and community-acquired gastroenteritis. Most cases of L hongkongensis infections occur in eastern countries. Information is lacking on the usefulness of matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI–TOF MS) for the identification of bacteria important in eastern countries. Using the Bruker database extended with 21 L hongkongensis reference strains, all 240 L hongkongensis isolates recovered from patients, fish, frogs and water were correctly identified, with 224 (93.3%) strains having top match scores ≥2.0. Notably, the strain of Chromobacterium violaceum was not reliably identified although it is included in the database. MALDI–TOF MS is useful for the accurate routine identification of L hongkongensis after adding reference L hongkongensis main spectra to the database. The number of strains for each species in MALDI–TOF MS databases should be expanded to cover intraspecies variability.