Characterization of fungal spores by laser desorption/ionization time-of-flight mass spectrometry.

A considerable volume of research has now been completed on the application of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to the analysis of bacteria; however, to date no definitive studies have been made using this technique on fungi. Preliminary studies on the application of the MALDI-MS methodology, previously developed for the analysis of bacteria, to the analysis of intact fungal spores are described here. MALDI-MS and electrospray mass spectrometry enable the high molecular weight analysis of proteins, glycoproteins, oligosaccharides and oligonucleotides. Using MALDI-MS with bacteria has demonstrated the ability to produce 'fingerprints' of the intact cells with the ions observed being associated with the proteinaceous components of the cell wall. This paper reports the adaptation of this technique to the direct analysis of fungal cells. The high percentage of carbohydrate in the fungal cell wall indicates that the ions observed in the mass spectrometric experiments may be of carbohydrate origin. Penicillium spp., Scytalidium dimidiatum and Trichophyton rubrum have been studied in this preliminary investigation and all show individually distinctive spectra which would appear to provide a profile of the cellular material with discrete peaks being observed over the mass range 2 to 13 kDa. The spectra obtained are reproducible within the method used but, as shown in our previous studies on bacteria, washing may selectively release components from the fungal cell wall.

The effect of solvent and matrix combinations on the analysis of bacteria by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

The ability to rapidly identify the taxonomic class of the wide variety of microorganisms involved in human and animal disease is becoming increasingly important, especially with the increasing development of resistance to the antibiotics which form the main defence against them. A number of groups have recognised the utility of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry MALDI-TOF in the analysis of these microorganisms. However, no consistent methodology has been developed which is in general use. In particular the use of different solvent extraction systems and mass spectrometric matrices can have significant effects on the quality of the data obtained. We have now studied a number of the commonly used matrices and a range of solvent systems of widely varying polarity in an attempt to devise an optimum analytical strategy for the rapid characterisation of these organisms by MALDI-TOFMS. The E. coli ATCC 9637 organisms were initially washed to remove growth medium contaminants, followed by extraction with one of a range of solvents prior to admixing with a number of different single matrices or binary and ternary combinations of these matrices. The results obtained indicate that a binary combination of 2-(4-hydroxyphenylazo)benzoic acid and 2-mercaptobenzothiazole (1:1) as matrix provides the best data after the proteinaceous material from the organism cell surface was extracted with 17% formic acid, 33% isopropyl alcohol and 50% water, (solvent 2 in this work).

Highly virulent pathogens--a post antibiotic era?

The spectrum of infectious diseases is changing rapidly. Emerging infectious agents present an intriguing constellation of nosocomial challenges. Antimicrobial resistance results in increased morbidity, mortality and costs of health care. Resistance to antimicrobial agents has been recorded since 1940 with penicillin resistant Escherichia coli (E coli) (Abraham and Chain 1940). A similar pencillin resistance was reported in 1944 in Staphylococus aureus (S. aureus) (Kirby 1944) Even before the widespread global use of penicillin, resistance had already been detected in both gram-positive and gram-negative organisms. The 1990s herald the era of multiple drug resistance. To grasp further the enormity and complexity of our modern antimicrobial resistance problem, one only needs to think about how many--how fast--and in how many settings (hospitals, clinics, outpatients nursing and long term facilities, etc), these pathogens have developed antimicrobial resistance: Multiple drug-resistant Mycobacterium tuberculosis, penicillin-resistant Streptococcus pneumonia, fluconzole-resistant Candida, methicillin-resistant S. aureus (MRSA), vancomycin-resistant Enterococci (VRE) and now S. aureus with reduced susceptibility to vancomycin. Given the dramatic increase in the incidence of multiple drug-resistant organisms--and now--the mounting evidence of resistance transfer from one organism to another, we will certainly witness a combined growth of nosocomial pathogens, for which there are no antibiotic solutions. Appropriate infection control measures for such resistant strains depend, in part, on the mechanisms of genetic information exchanged among micro-organisms. Clearly we need to strengthen the basic tenets of infection prevention and control; hygiene, engineering and microbial barriers, to prevent transinfection. We need to control horizontal nosocomial transmission of organisms. Contaminated environmental surfaces are a reservoir for resistant organisms such as MRSA (Boyce et al 1997) and VRE (Karanfil et al 1992). Stringent infection control policies need to be developed and implemented. A comprehensively applied infection control programme will reduce the dissemination of resistant strains. Each patient care setting must examine its current practices and review the outcome efficacy. A consensus development conference to develop centres for disease control (CDC) formal guidelines against vancomycin intermediate-level resistant staphylococcus aureus (VISA) and vancomycin-resistant staphylococcus aureus (VRSA) may take a year or more to convene. This paper will examine the basic considerations currently offered by the CDC which may be valuable starting points for the enhancement of current infection control practices. Perspectives of the Society for Healthcare Epidemiology of America (SHEA) will also be included.

The characterization of micro-organisms by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

A variety of gram-positive and gram-negative intact bacterial cells have been analysed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and shown to provide fingerprint mass spectra with discrete peaks being observed over the mass range from 3 to 40 kDa. The spectra show both more peaks and peaks at a higher mass/charge ratio than have hitherto been reported for these micro-organisms and would appear to provide a profile of cellular proteinaceous material. The spectra are shown to be reproducible over variable time periods of up to three months and factors affecting reproducibility are discussed. The procedure, which requires minimal sample preparation, yields results in 30-40 minutes and allows visual identification of species- and strain-specific biomarkers for the characterization of the organisms. The importance of accurately defining sample preparation methodologies is central to the ability of the technique to generate reliable and reproducible data.

Induction of amenorrhea during hormone replacement therapy: optimal micronized progesterone dose. A multicenter study.

The effects of oral micronized progesterone on the endometrium and bleeding pattern have been assessed in a multicenter study of 101 postmenopausal patients. During a minimum of 6 cycles, the participants received either percutaneous 17 beta-estradiol (1.5 mg/day) associated with micronized progesterone (100 mg/day), given at bedtime for 21/28 days or 25 days/calendar month (n = 98) [1], or E2 (3 mg/day) for 25 days associated with progesterone (300 mg/day), from day 16 to day 25 (n = 3) [2], according to their willingness to induce, or not, cyclic withdrawal bleeding. Each endometrial biopsy performed at 6-month minimum was assessed by two independent pathologists: results showed 61% quiescent without mitosis, 23% mildly active with very rare mitoses and 8% partial secretory endometrium. The remaining biopsies showed inadequate tissue (4%) or a sub-atrophy (4%). No hyperplasia was found by any pathologist. In the case of inadequate material, the mean thickness of endometrial mucosa measured by ultrasonography was 3.9 mm. Amenorrhea incidence was 93.3 and 91.6% at the 3rd and 6th month of therapy, respectively. No bleeding occurred in more than 80% of women. The results show that a low dose of oral progesterone (100 mg/day), given during 25 days, efficiently protects the endometrium by fully inhibiting mitoses and induces amenorrhea in the majority of postmenopausal women, allowing better compliance to long-term therapy.

OSHA's final rule on occupational exposure to bloodborne pathogens.

This report is intended as a brief review of some of the key points of the new OSHA rule. It should not be used in place of the complete rule, and you should contact your legal advisor with any questions about interpretations of the rule. Because circumstances of occupational exposure vary among employers, it is advisable that employers be thoroughly acquainted with all requirements of the rule. The preamble to the rule, starting on page 64004 of the December 6, 1991 Federal Register, may provide helpful background information for understanding specific provisions of the rule.

Safety precautions for laser surgery.

This article discusses the dangers of laser surgery and how to best guard against them by using the appropriate products and protective gear and by setting up a laser safety committee to continuously address laser safety problems.

Laser surgery--the future is here.

Laser surgery experts have indicated that surgical lasers provide a number of potential benefits when compared to conventional surgical methods. The most significant are: a reduction in hospital costs; an opportunity to increase the surgical case load; and the opportunity to provide competitive, "high-tech" clinical services to the hospital's community. Lasers change the procedures utilized in the hospital in terms of procurement, processing, safety and other areas. It is incumbent upon materials managers and CS personnel to become familiar with and keep pace with these changes. As a these provider, it is important that you follow the laser revolution. The emergence of lasers in surgery will affect almost every segment of hospital operations, and each department manager can turn change into opportunity for their facility.