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Antifungal susceptibilities of non-Aspergillus filamentous fungal pathogens cannot always be inferred from their identification. Here we determined, using the Sensititre(®) YeastOne(®) YO10 panel, the in vitro activities of nine antifungal agents against 52 clinical isolates of emergent non-Aspergillus moulds representing 17 fungal groups in Australia. Isolates comprised Mucorales (n = 14), Scedosporium/Lomentospora spp. (n = 18) and a range of hyaline hyphomycetes (n = 9) and other dematiaceous fungi (n = 11). Excluding Verruconis gallopava, echinocandins demonstrated poor activity (MICs generally >8 mg/L) against these moulds. Lomentospora prolificans (n = 4) and Fusarium spp. (n = 6) demonstrated raised MICs to all antifungal drugs tested, with the lowest being to voriconazole and amphotericin B (AmB), respectively (geometric mean MICs of 3.4 mg/L and 2.2 mg/L, respectively). All Scedosporium apiospermum complex isolates (n = 14) were inhibited by voriconazole concentrations of ≤0.25 mg/L, followed by posaconazole and itraconazole at ≤1 mg/L. Posaconazole and AmB were the most active agents against the Mucorales, with MIC90 values of 1 mg/L and 2 mg/L, respectively, for Rhizopus spp. For dematiaceous fungi, all isolates were inhibited by itraconazole and posaconazole concentrations of ≤0.5 mg/L (MIC90, 0.12 mg/L and 0.25 mg/L, respectively), but voriconazole and AmB also had in vitro activity (MIC90, 0.5 mg/L and 1 mg/L, respectively). Differences in antifungal susceptibility within species and between species within genera support the need for testing individual patient isolates to guide therapy. The Sensititre(®) YeastOne(®) offers a practical alternative to the reference methodology for susceptibility testing of moulds.

Mucormycosis is the second most common cause of invasive mould infection and causes disease in diverse hosts, including those who are immuno-competent. We conducted a multicentre retrospective study of proven and probable cases of mucormycosis diagnosed between 2004-2012 to determine the epidemiology and outcome determinants in Australia. Seventy-four cases were identified (63 proven, 11 probable). The majority (54.1%) were caused by Rhizopus spp. Patients who sustained trauma were more likely to have non-Rhizopus infections relative to patients without trauma (OR 9.0, p 0.001, 95% CI 2.1-42.8). Haematological malignancy (48.6%), chemotherapy (42.9%), corticosteroids (52.7%), diabetes mellitus (27%) and trauma (22.9%) were the most common co-morbidities or risk factors. Rheumatological/autoimmune disorders occurred in nine (12.1%) instances. Eight (10.8%) cases had no underlying co-morbidity and were more likely to have associated trauma (7/8; 87.5% versus 10/66; 15.2%; p

Central nervous system (CNS) cryptococcosis is most commonly encountered among HIV-infected and other immunosuppressed hosts but is less well-characterized among non-immunosuppressed patients.We conducted a three year, prospective, observational study to compare the clinical manifestations and outcome of CNS cryptococcosis in three patient populations: HIV-infected patients (n = 54), HIV-negative immunosuppressed patients (n = 21), and non-immunosuppressed patients (n = 11).Time from initial symptoms to presentation did not differ between the groups. HIV-infected patients were significantly more likely to present with fevers (p 0.0001), but were less likely to have abnormal mental status, CNS mass lesions and pulmonary involvement (p = 0.001, 0.01 and 0.03, respectively). The clinical manifestations among HIV-negative immunosuppressed patients were generally intermediate to the other groups. Overall, acuity of illness was worse among non-immunosuppressed patients, as measured by APACHE II scores (p = 0.02). Intracranial pressure was higher in HIV-infected and non-immunosuppressed patients than immunosuppressed patients (p = 0.008 and 0.01, respectively). Repeated lumbar punctures were more common among HIV-infected patients (p = 0.01). There was a trend toward more frequent placement of permanent CNS shunts among non-HIV patients (p = 0.05). The mortality rate was greatest for non-immunosuppressed patients (p = 0.04).CNS cryptococcosis in non-immunosuppressed patients was associated with poorer prognosis. Our findings suggest that host immune responses may contribute to pathogenesis of CNS cryptococcosis, with more intact immune function associated with increased CNS-related morbidity and overall mortality.

First-aid education for the management of burns advocates cool running water over burnt skin to limit soft tissue damage. However, the water used may itself constitute a risk. We report three cases of severe invasive and necrotizing infection in patients who used or immersed themselves in contaminated water in an attempt to extinguish the fire following acute major burns. Wound cultures from all patients yielded Aeromonas hydrophila and two yielded Bacillus cereus. One patient had a complex polymicrobial infection, including zygomycosis with Rhizomucor variabilis. All patients were treated aggressively with wound débridement, including one patient who required bilateral lower limb amputations to control progressive infection. All infections were successfully treated and all patients survived their burn injuries. We review the management of burns complicated by exposure to contaminated water leading to burn wound infections. We describe commonly reported organisms from various water sources, the appropriate initial empirical antimicrobial chemotherapy and present the clinician with a proposed algorithm for managing these serious infections.

We report the development and evaluation of a real-time PCR assay using the LightCycler instrument for the detection of C. albicans and A. fumigatus DNA in whole blood. Recently published consensus criteria for the diagnosis of invasive fungal infection (IFI) were used for all patient samples. Unique and published primer pairs were developed and assessed for sensitivity, specificity, and reproducibility to detect C. albicans and A. fumigatus DNA in samples spiked with purified DNA, and whole blood samples from 8 high-risk patients and 45 negative controls. The real-time assay demonstrated an analytical sensitivity of 10 fg of purified C. albicans and A. fumigatus DNA and was found to be specific for each species. The standardized approach was highly reproducible and detected C. albicans and A. fumigatus DNA in two patients with proven IFI and in one patient with a possible IFI. In addition, we report for the first time the use of recently published international consensus criteria for the diagnosis of IFI in the evaluation of a mildly invasive fungal diagnostic assay. Standardized clinical criteria and a more standardized approach to detect fungal DNA in less invasive patient samples, may permit a more reliable comparison of future studies. A rapid real-time detection of fungal DNA in whole blood, combined with standard clinical markers of response, may be more useful for monitoring patients at risk of developing IFI than other diagnostic methods currently available.

A quantitative real-time PCR targeting the Pneumolysin (ply) gene of Streptococcus pneumoniae was developed for the LightCycler instrument using Fluorescence Resonance Energy Transfer (FRET) probes. All common S. pneumoniae serotypes were detected while other bacteria and viruses were not. The sensitivity was determined to be between one and ten target copies per reaction. The PCR was applied to six CSF and 16 whole blood specimens from 17 patients with laboratory proven invasive pneumococcal disease. One hundred percent of CSF specimens and 69% of whole blood specimens were PCR positive. The bacterial loads were determined to be 7.6 to 6.01 x 10(5) copies/microL for the six CSF specimens, and 0.08 to 5.4 x 10(2) copies/microL for the 16 whole blood specimens. Ninety-seven percent of 30 culture and Gram's stain negative CSF specimens and 100% of 50 normal whole blood specimens were PCR negative. This highly sensitive and specific PCR assay has the potential to provide sufficiently rapid results to improve antibiotic treatment of S.pneumoniae infections, while bacterial load quantitation has opened up exciting possibilities for patient management.

The last few decades have seen significant changes in health care with increasing numbers of heavily immunocompromised patients. Because of new and more aggressive treatments, patients with severe immune defects are surviving longer and the spectrum of fungal pathogens is increasing. Until relatively recently, Candida albicans and Aspergillus fumigatus were considered the only important nosocomial fungal pathogens. However, non-albicans Candida species and other yeasts, together with an increasing range of moulds apart from A. fumigatus are now reported to cause nosocomial fungal infections. Candidaemia is the fourth most frequent cause of health care associated bloodstream infection in the USA. Candidaemia has a crude mortality of40-80%, and an attributable mortality of approximately 40%. Australian data, although scanty, suggest that invasive fungal infections (IFIs) are also a significant cause of health care associated infections in Australia. The widespread use of azoles for prophylaxis and treatment has been linked with the emergence of non-albicans Candida species and other yeasts. A. fumigatus and a number of other moulds are also important nosocomial pathogens, associated with both prolonged lengths of hospital stay and very high crude mortality rates. Therefore prevention of nosocomial fungal infections is an increasingly important aspect of infection control. Both old and new infection control strategies will need to be implemented to protect our highly immunosuppressed patients particularly from this increasing noscomial threat.

The epidemiology of invasive fungal disease (IFD) due to filamentous fungi other than Aspergillus may be changing. We analysed clinical, microbiological and outcome data in Australian patients to determine the predisposing factors and identify determinants of mortality. Proven and probable non-Aspergillus mould infections (defined according to modified European Organization for Research and Treatment of Cancer/Mycoses Study Group criteria) from 2004 to 2012 were evaluated in a multicentre study. Variables associated with infection and mortality were determined. Of 162 episodes of non-Aspergillus IFD, 145 (89.5%) were proven infections and 17 (10.5%) were probable infections. The pathogens included 29 fungal species/species complexes; mucormycetes (45.7%) and Scedosporium species (33.3%) were most common. The commonest comorbidities were haematological malignancies (HMs) (46.3%) diabetes mellitus (23.5%), and chronic pulmonary disease (16%); antecedent trauma was present in 21% of cases. Twenty-five (15.4%) patients had no immunocompromised status or comorbidity, and were more likely to have acquired infection following major trauma (p

A previously well 42-year-old man presented with 4 months of intermittent short-term memory loss, fever, headache, and 20 kg weight loss. He was febrile (38°C), abulic, and disorientated with fl uctuating alertness requiring frequent prompting and supervision to do all selfcare tasks. He had impaired hand dexterity, mild midline ataxia, and subtle neck stiff ness. The provisional diagnosis was an encephalopathy and antibiotics were deferred. Initial cerebrospinal fl uid (CSF) was turbid, with 1340×106 cells per L (95% mononuclear cells) and protein concentration of 11·0 g/L; cultures were negative. White cell count was 9800×106 cells per L and C-reactive protein concentration was less than 1 mg/L. Cranial CT scan with contrast was normal, and blood cultures and repeat lumbar puncture were sterile. MRI suggested granulomatous infl ammatory or non-infective processes—for example, tuberculous or carcinomatous meningitis, respectively (fi gure). A third lumbar puncture (15 days after the fi rst) showed 793×106 cells per L (55% mononuclear cells), protein concentration of 5·5 g/L, and glucose concentration of 1·5 mmol/L (CSF/ serum ratio of 0·26). CSF Gram stain showed no bacteria, but cultures yielded Neisseria meningitidis. Intravenous ceftriaxone 2 g twice-daily was given, which improved the patient’s cognitive and neurological status within days. Lumbar puncture and MRI done 6 weeks postdischarge were both normal. Retrospective analysis of CSF samples by PCR, confi rmed N meningitidis serogroup B DNA from all CSF specimens, and identical variable region PorA (P1·22,14) genotypes. Further investigations revealed a low serum mannose binding lectin (MBL) concentration. The patient remains healthy 24 months after discharge without neurological sequelae. N meningitidis infection rarely presents as chronic meningitis or encephalopathy. Since all initial specimens were sterile, it is likely that the patient’s CSF had a low number of bacteria. Current concepts in bacterial pathogenesis suggest that when microbial population densities are reduced, quorum-sensing systems down regulate the bacterial virulence genes, thereby partly avoiding host detection. This, together with the patient’s MBL defi ciency, may explain the indolent presentation of this normally virulent organism.