Susceptibility testing of fluconazole by the NCCLS broth macrodilution method, E-test, and disk diffusion for application in the routine laboratory

Antifungal susceptibility testing may be an important aid in the treatment of patients with life-threatening yeast infections. In order to establish the suitability of different susceptibility test methods for fluconazole with yeasts, the Rosco tablet and the E-test were compared with the gold standard NCCLS broth macrodilution method for 106 yeast strains. These included 102 clinical isolates of Candida spp., including Candida glabrata (n = 30), Candida albicans (n = 20), Candida tropicalis (n = 13), Candida parapsilosis (n = 10), Candida krusei (n = 8), plus Cryptococcus neoformans (n = 3), Saccharomyces cerevisiae (n = 2), and 16 strains belonging to other Candida spp. Four American Type Culture Collection strains of Candida were included as quality controls. The NCCLS method was found to be too complex and labor-intensive for routine testing. The E-test is an accurate alternative, but experience in determining MICs and careful attention to procedural details are critically important. The Rosco tablet showed the best agreement with the NCCLS reference method, especially when newly established breakpoints of R < or = 10 mm and S > or = 21 mm were used.     

Bacteremia due to Moraxella atlantae in a cancer patient

A gram-negative alkaline phosphatase- and pyrrolidone peptidase-positive rod-shaped bacterium (CCUG 45702) was isolated from two aerobic blood cultures from a female cancer patient. No identification could be reached using phenotypic techniques. Amplification of the tRNA intergenic spacers revealed fragments with lengths of 116, 133, and 270 bp, but no such pattern was present in our reference library. Sequencing of the 16S rRNA gene revealed its identity as Moraxella atlantae, a species isolated only rarely and published only once as causing infection. In retrospect, the phenotypic characteristics fit the identification as M. atlantae (formerly known as CDC group M-3). Comparative 16S rRNA sequence analysis indicates that M. atlantae, M. lincolnii, and M. osloensis might constitute three separate genera within the MORAXELLACEAE: After treatment with amoxicillin-clavulanic acid for 2 days, fever subsided and the patient was dismissed.     

Isolation of Moraxella canis from an ulcerated metastatic lymph node

Moraxella canis was isolated in large numbers from an ulcerated supraclavicular lymph node of a terminal patient, who died a few days later. Although the patient presented with septic symptoms and with a heavy growth of gram-negative diplococci in the lymph node, blood cultures remained negative. M. canis is an upper-airway commensal from dogs and cats and is considered nonpathogenic for humans, although this is the third reported human isolate of this species.     

Selective medium for Branhamella catarrhalis with acetazolamide as a specific inhibitor of Neisseria spp.

Several semiselective media for Branhamella catarrhalis have been proposed. These media allow growth of all members of the family Neisseriaceae, and further differentiation is necessary. By addition of 10 micrograms of acetazolamide, a carbonic anhydrase inhibitor, per ml and incubation in air, a medium was created which reduced growth of Neisseria spp. When saliva samples from 178 healthy schoolchildren were screened for the presence of B. catarrhalis, the carrier rate for this organism was estimated to be 48.9% with the selective medium compared with 12.4% when a semiselective medium, which contains only 10 micrograms of vancomycin, 5 micrograms of trimethoprim, and 2 micrograms of amphotericin B per ml, was used and 6.2% when a nonselective blood agar plate was used. The number of Neisseria spp. isolated dropped from 297 on the semiselective agar to 55 on the selective agar.     

Application of tRNA intergenic spacer PCR for identification of Enterococcus species

tRNA intergenic spacer PCR (tDNA-PCR) was evaluated for its usefulness in the differentiation of enterococcal species of human and animal origin. This technique was carried out for 124 strains belonging to 17 enterococcal species and generated DNA fragments, which were separated by capillary electrophoresis. tDNA-PCR enabled us to discriminate for all species tested. Enterococcus faecium showed minor but reproducible differences with Enterococcus durans, while Enterococcus hirae was easily distinguishable. Enterococcus avium, Enterococcus malodoratus, and Enterococcus raffinosus generated highly similar though distinctive patterns.     

Antibiotic susceptibility of Atopobium vaginae

Background  

Previous studies have indicated that a recently described anaerobic bacterium, Atopobium vaginae is associated with bacterial vaginosis (BV). Thus far the four isolates of this fastidious micro-organism were found to be highly resistant to metronidazole and susceptible for clindamycin, two antibiotics preferred for the treatment of BV.     

New Insights into Understanding the Mechanisms,Pathogenesis, and Management of Malignant Mesotheliomas

Malignant mesothelioma (MM) is a relatively rare but devastating tumor that is increasing worldwide. Yet, because of difficulties in early diagnosis and resistance to conventional therapies, MM remains a challenge for pathologists and clinicians to treat. In recent years, much has been revealed regarding the mechanisms of interactions of pathogenic fibers with mesothelial cells, crucial signaling pathways, and genetic and epigenetic events that may occur during the pathogenesis of these unusual, pleiomorphic tumors. These observations support a scenario whereby mesothelial cells undergo a series of chronic injury, inflammation, and proliferation in the long latency period of MM development that may be perpetuated by durable fibers, the tumor microenvironment, and inflammatory stimuli. One culprit in sustained inflammation is the activated inflammasome, a component of macrophages or mesothelial cells that leads to production of chemotactic, growth-promoting, and angiogenic cytokines. This information has been vital to designing novel therapeutic approaches for patients with MM that focus on immunotherapy, targeting growth factor receptors and pathways, overcoming resistance to apoptosis, and modifying epigenetic changes.CME Accreditation Statement: This activity (“ASIP 2013 AJP CME Program in Pathogenesis”) has been planned and implemented in accordance with the Essential Areas and policies of the Accreditation Council for Continuing Medical Education (ACCME) through the joint sponsorship of the American Society for Clinical Pathology (ASCP) and the American Society for Investigative Pathology (ASIP). ASCP is accredited by the ACCME to provide continuing medical education for physicians.The ASCP designates this journal-based CME activity (“ASIP 2013 AJP CME Program in Pathogenesis”) for a maximum of 48 AMA PRA Category 1 Credit(s)™. Physicians should only claim credit commensurate with the extent of their participation in the activity.CME Disclosures: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose.Malignant mesotheliomas (MMs), among the most aggressive tumors, arise most often from the mesothelial cells that line the pleura, peritoneum, and, occasionally, the pericardium. Because of the multifaceted properties of mesothelium that maintain a protective barrier but also produce components of the extracellular matrix, hyaluronan and other lubricants, chemokines and cytokines, and fibrinolytic and procoagulant factors, understanding its complex biology is a challenge. The intermediate filament pattern of mesothelial cells, suggesting an epithelial–mesodermal hybrid morphology, and their several patterns of differentiation during the neoplastic process suggest their transformation to malignancy is complicated and raises the question of whether one is studying a single tumor type or multiple subgroups of tumors.MMs are most commonly attributed to occupational exposures to asbestos, a regulatory term for a group of fibrous silicates that occur as needle-like amphiboles (crocidolite, amosite tremolite, anthophyllite, and antigorite) or curly serpentine (chrysotile) fibers. Although each of these fibers has its own distinctive properties, the fibrous nature and biopersistance of these inhaled fibers may be key to carcinogenic events that occur during the long latency periods (mean, 30 to 45 years) of most MMs. Most intensely investigated are chrysotile, the most commonly used type of asbestos historically (>90% use worldwide), and crocidolite, the asbestos type associated most often with MMs in humans^1,2 (Figures 1 and 2). The morphology of crocidolite asbestos is similar to nonasbestos fibers of erionite or Libby amphibole, other naturally occurring minerals associated with the development of MMs.^5,6 However, 20% to 25% of individuals with MM have no documented exposure to asbestos or other fibers, suggesting familial susceptibility (sporadic or idiopathic MM), unknown exposure to in-place or naturally occurring asbestos, or other causative agents, such as chemicals, radiation, and viruses.⁷Open in a separate windowFigure 1Properties of chrysotile (white) asbestos. A: Image of bundle of curly chrysotile fibers before processing. B: Scanning electron micrograph of chrysotile fibers (arrows) causing deformation of red blood cells. Chrysotile is positively charged, hemolytic, and cytolytic, primarily due to its magnesium content. Leaching of magnesium renders chrysotile less toxic and also results in chrysotile fiber dissolution over time. C: Scanning electron micrograph of interaction of long chrysotile fiber with the respiratory epithelium of the alveolar duct junction after inhalation by rats. Arrowheads show points of contact with and between epithelial cells. Subsequent penetration into and between cells leads to fiber deposition in the lung interstitum and access to the visceral pleura and pleural space. D: Polarized microscopy showing chrysotile fibers and fibrils.Photomicrograph is a courtesy of Lee Poye (J3 Resources, Inc., Houston, TX) Original magnification, ×100.Open in a separate windowFigure 2Properties of crocidolite, or blue, asbestos. A: Riebeckito ore showing veins of crocidolite asbestos fibers (arrow) before processing. B: Scanning electron micrograph showing morphology of needle-like fibers. C: Early penetration of a crocidolite fiber into the differentiated tracheobronchial epithelium in tracheal organ culture. D: Growth of metaplastic cells over long fibers of crocidolite observed at 1 month in this model.³ These events have not been captured in the pleura in animal inhalation models or in clinical specimens in humans, but mesothelial cells undergo proliferation, as measured by cell counts, or immunochemical markers have been observed in response to crocidolite asbestos in vitro and after inhalation by rats.⁴Because asbestos fibers neither appear to be metabolized nor directly interact with DNA, they are unlike most chemical carcinogens. The sensitivity of human mesothelial cells to fibers of high aspect (length to diameter) ratio is also perplexing, as are the phenomena governing fiber transport to the parietal pleura where most MMs are thought to develop. Although much insight exists on understanding how fibers (particularly high iron-containing amphibole asbestos types) generate reactive oxygen and nitrogen species to induce inflammation and cell signaling pathways important in proliferation and transformation, how these cellular events converge in the pathogenesis of MM remains enigmatic. This review amalgamates current observations in the field and their implications in strategies to prevent and manage MMs in patients.     

Gynaecological cancers in pregnancy

Cervical and ovarian cancers are the most common gynaecological cancers diagnosed during pregnancy. In early-stage cervical cancer during the first and at the beginning of the second trimester, the two main considerations for management of the patient are the tumour size (and stage) and nodal staging. MRI and laparoscopic lymphadenectomy are useful for clinicians planning a potentially conservative approach. The management of patients with locally advanced cervical disease is controversial and should be discussed on a case-by-case basis according to the tumour size, radiological findings, the term of pregnancy, and the patient's wishes. Different histological types of malignant ovarian diseases arise during pregnancy and their management depends on the diagnosis (histological subtypes, tumour differentiation, and nodal status), the tumour stage, and the trimester of the pregnancy. In patients with peritoneal spread or high-risk early-stage disease, neoadjuvant chemotherapy with pregnancy preservation could be appropriate.     

Quantification of the breakpoint cluster region rearrangement for clinical monitoring in Philadelphia chromosome-positive chronic myeloid leukemia

The purpose of this report was to evaluate scintigraphy analysis of Southern blot hybridization as a method to quantify the breakpoint cluster region (BCR) rearrangement of Philadelphia chromosome (Ph)+ chronic myelogenous leukemia (CML). Cytogenetic and molecular studies performed simultaneously on 474 bone marrow and/or blood samples from 300 patients treated with alpha-interferon-based therapy were compared. Molecular results were expressed as the percentage of rearranged BCR bands versus the total scintigraphic signal. The percentage of Ph+ metaphases was calculated on 25 metaphases. The results of molecular studies obtained on both peripheral blood and bone marrow samples were identical. The rank correlation between the BCR quantification and the percentage of Ph positivity in 465 samples was excellent (r = .78). However, of 99 samples with a normal karyotype, 24% had a BCR rearrangement. Of 86 samples with no BCR rearrangement, 13% showed a Ph chromosome. Of 49 samples with partial cytogenetic remission (Ph+ metaphases, 1% to 34%), 23% had no BCR rearrangement. In samples with a minor or no cytogenetic response (Ph+ metaphases, > 34%), BCR analysis overestimated the degree of response in 73 of 326 samples (22%). Nevertheless, survival analysis by BCR quantification level showed statistically better outcome for patients in complete or partial molecular response (P < .01). Molecular quantification of BCR was useful in monitoring the course of Ph+ CML. This method, which can be used on peripheral blood, detected residual disease not shown by cytogenetic analysis and was prognostically relevant as a measure of disease suppression.