The impact of anti-mould prophylaxis on Aspergillus PCR blood testing for the diagnosis of invasive aspergillosis.

BACKGROUND: The performance of the galactomannan enzyme immunoassay (GM-EIA) is impaired in patients receiving mould-active antifungal therapy. The impact of mould-active antifungal therapy on Aspergillus PCR testing needs to be determined. OBJECTIVES: To determine the influence of anti-mould prophylaxis (AMP) on the performance of PCR blood testing to aid the diagnosis of proven/probable invasive aspergillosis (IA). METHODS: As part of the systematic review and meta-analysis of 22 cohort studies investigating Aspergillus PCR blood testing in 2912 patients at risk of IA, subgroup analysis was performed to determine the impact of AMP on the accuracy of Aspergillus PCR. The incidence of IA was calculated in patients receiving and not receiving AMP. The impact of two different positivity thresholds (requiring either a single PCR positive test result or ≥2 consecutive PCR positive test results) on accuracy was evaluated. Meta-analytical pooling of sensitivity and specificity was performed by logistic mixed-model regression. RESULTS: In total, 1661 (57%) patients received prophylaxis. The incidence of IA was 14.2%, significantly lower in the prophylaxis group (11%-12%) compared with the non-prophylaxis group (18%-19%) (P < 0.001). The use of AMP did not affect sensitivity, but significantly decreased specificity [single PCR positive result threshold: 26% reduction (P = 0.005); ≥2 consecutive PCR positive results threshold: 12% reduction (P = 0.019)]. CONCLUSIONS: Contrary to its influence on GM-EIA, AMP significantly decreases Aspergillus PCR specificity, without affecting sensitivity, possibly as a consequence of AMP limiting the clinical progression of IA and/or leading to false-negative GM-EIA results, preventing the classification of probable IA using the EORTC/MSGERC definitions.

Tuberculosis incidence in the Irish Traveller population in Ireland from 2002 to 2013.

The health status of the Irish Traveller ethnic minority is low compared to the general population in Ireland in terms of infant mortality rates and life expectancies. Respiratory disease is an area of health disparity manifested as excess mortalities in Traveller males and females. In this study, we examined the available data with regard to tuberculosis (TB) notifications in Ireland from 2002 to 2013. We found an increase in TB notifications in Irish Travellers from 2010 onwards. This resulted in a crude incidence rate for TB in Irish Travellers that was approximately threefold higher than that of the white Irish-born population in 2011 and 2012. An outbreak of TB in Irish Travellers in 2013 increased this differential further, but when outbreak-linked cases were excluded, a higher incidence rate was still observed in Irish Travellers relative to the general population and to white Irish-born. The mean age of a TB patient was 26 years in Irish Travellers compared to 43 years in the general population, and 49 years in white Irish-born. Based on available data, Irish Travellers exhibit a higher incidence rate and younger age distribution of TB compared to white Irish-born and the general population. These observations emphasize the importance of routine use of ethnicity identifiers in the management of TB and other notifiable communicable illnesses in Ireland. They also have implications for the orientation of preventive services to address health disparities in Irish Travellers and other ethnic minority groups.

Multicenter comparison of serum and whole-blood specimens for detection of Aspergillus DNA in high-risk hematological patients.

Samples from patients at high risk for invasive aspergillosis (IA) were prospectively collected and analyzed for the presence of molecular markers of fungal infection. Serum specimens were screened for galactomannan and Aspergillus DNA, and whole-blood specimens were screened only for Aspergillus DNA. Fungal infections were categorized according to the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group, National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria. Forty-seven cases (proven and probable IA) and 31 controls (no evidence of IA) were selected retrospectively for this case-control study, comprising 803 samples, in order to determine the performance of whole-blood PCR, serum PCR, and serum galactomannan testing. Although no single assay was able to detect every case of IA, a combination of different assays provided the best performance. There was no significant difference between the use of whole-blood and serum specimens for PCR-based diagnosis of IA, but there was a trend for whole blood to be more sensitive (85% versus 79%) and to yield an earlier positive result (36 days versus 15 days) than for serum. However, DNA extraction from serum specimens is easier and faster than that from whole-blood specimens, and it allows the same specimen to be used for both galactomannan and PCR assays. In conclusion, the appropriate sample type for DNA extraction should be determined by the local requirements and the technical platforms available at each individual center. A combination of biomarker tests offered the best diagnostic utility for detecting IA.

A snapshot of genetic lineages of Mycobacterium tuberculosis in Ireland over a two-year period, 2010 and 2011.

Mycobacterial interspersed repetitive-unit-variable-number tandem repeat typing alone was used to investigate the genetic lineages among 361 Mycobacterium tuberculosis strains circulating in Ireland over a two-year period, 2010 and 2011. The majority of isolates, 63% (229/361), belonged to lineage 4 (Euro-American), while lineages 1 (Indo-Oceanic), 2 (East-Asian) and 3 (East-African­Indian) represented 12% of isolates each (42/361, 45/361, and 45/361, respectively). Sub-lineages Beijing (lineage 2), East-African­Indian (lineage 1) and Delhi/central-Asian (lineage 3) predominated among foreign-born cases, while a higher proportion of Euro-American lineages were identified among cases born in Ireland. Eighteen molecular clusters involving 63 tuberculosis (TB) cases were identified across four sub-lineages of lineage 4. While the mean cluster size was 3.5 TB cases, the largest cluster (involving 12 Irish-born cases) was identified in the Latin American­Mediterranean sub-lineage. Clustering of isolates was higher among Irish-born TB cases (47 of 63 clustered cases), whereas only one cluster (3/63) involved solely foreign-born individuals. Four multidrug-resistant cases identified during this period represented lineages 2 and 4. This study provides the first insight into the structure of the M. tuberculosis population in Ireland.

The largest prison outbreak of TB in Western Europe investigated using whole-genome sequencing.

BACKGROUND: In March 2011, the Department of Public Health East in Ireland were notified of two cases of TB in two prisoners sharing a cell. We define the resulting outbreak and highlight the role of public health and laboratory-based molecular epidemiology in mapping and control of a prison outbreak.METHODS: Cases were identified through clinical presentation, contact tracing, case-finding exercise or enhanced laboratory surveillance. Mycobacterium tuberculosis isolates were genotyped and underwent whole-genome sequencing (WGS).RESULTS: Of the 34 cases of TB linked to the outbreak, 27 were prisoners (79%), 4 prison officers (12%) and 3 community cases (9%). M. tuberculosis was isolated from 31 cases (culture positivity: 91%). A maximum of six single-nucleotide polymorphisms separated the isolates, with 22 being identical, suggestive of a highly infectious 'super-spreader´ within the prison. Isolates belonged to the Beijing sub-lineage, and were susceptible to first-line anti-TB agents. A case-finding exercise incidentally detected a prisoner with multidrug-resistant TB. Of the 143 prison officers screened, 52% had latent TB infection. Litigation costs exceeded five million euros.CONCLUSION: This constitutes the largest prison outbreak of TB in Western Europe investigated using WGS. A robust prison entry TB screening and education programme is required to effect better TB control, and prevent future outbreaks and attendant litigation.

Antifungal stewardship in critical care: Implementing a diagnostics-driven care pathway in the management of invasive candidiasis.

BACKGROUND: Invasive candidiasis (IC) is the most common invasive fungal disease in patients admitted to critical care and is associated with high mortality rates. Diagnosis can be delayed by the poor sensitivity of culture-based methods, leading to unnecessary use of empirical antifungal therapy (EAFT). The fungal biomarker (1-3)-ß-d-glucan (BDG) has been shown to aid in the diagnosis of IC in critical care and has been incorporated into antifungal stewardship (AFS) programmes. AIM: To describe our experience using a diagnostics-driven AFS programme incorporating the fungal biomarker BDG, analyse its impact on antifungal therapy (AFT), and gain an improved understanding of the epidemiology of IC in our critical care unit (CrCU). METHODS: An AFS care pathway incorporating BDG was introduced in the CrCU in St James's Hospital, Dublin. Following an educational programme, compliance with the pathway was prospectively audited between December 1st, 2017 and July 31st, 2018. RESULTS AND CONCLUSION: One hundred and nine AFT episodes were included, of which 95 (87%) had a BDG sent. Of those with BDG results available at the time of decision-making, 38 (63%) were managed in accordance with the care pathway. In compliant episodes without IC, median EAFT duration was 5.5 days [IQR 4-7] and no increase in mortality or subsequent IC was observed. Although adopting a diagnostics-driven approach was found to be useful in the cohort of patients with BDG results available, the use of once-weekly BDG testing did not result in an observed reduction in the consumption of anidulafungin, highlighting an important limitation of this approach.

Prevention of healthcare-associated invasive aspergillosis during hospital construction/renovation works.

The association between healthcare-associated invasive aspergillosis and hospital construction/building works is well recognized. This infection can cause significant morbidity and mortality and imposes a substantial burden on the healthcare system. The population of patients at risk for this opportunistic infection has expanded and multi-triazole drug resistance has emerged globally. Hence the need for a multi-faceted approach to prevent acquisition of invasive aspergillosis in acute care settings. This article is a summary of the Irish National Guidelines for the prevention of healthcare-associated aspergillosis which is based on published reports, international clinical guidelines, official engineering standards, and technical guidelines. We discuss the key recommendations and strategies for the prevention of invasive aspergillosis from the planning/pre-construction, construction, and post-construction phases. The importance of multi-disciplinary team involvement, education, and communication is emphasized.

Molecular epidemiology of multi- and extensively-drug-resistant Mycobacterium tuberculosis in Ireland, 2001-2014.

OBJECTIVES: The primary objective of this work was to examine the acquisition and spread of multi-drug resistant (MDR) tuberculosis (TB) in Ireland. METHODS: All available Mycobacterium tuberculosis complex (MTBC) isolates (n = 42), from MDR-TB cases diagnosed in Ireland between 2001 and 2014, were analysed using phenotypic drug-susceptibility testing, Mycobacterial-Interspersed-Repetitive-Units Variable-Number Tandem-Repeat (MIRU-VNTR) genotyping, and whole-genome sequencing (WGS). RESULTS: The lineage distribution of the MDR-TB isolates comprised 54.7% Euro-American, 33.3% East Asian, 7.2% East African Indian, and 4.8% Indo-Oceanic. A significant association was identified between the East Asian Beijing sub-lineage and the relative risk of an isolate being MDR. Over 75% of MDR-TB cases were confirmed in non-Irish born individuals and 7 MIRU-VNTR genotypes were identical to clusters in other European countries indicating cross-border spread of MDR-TB to Ireland. WGS data provided the first evidence in Ireland of in vivo microevolution of MTBC isolates from drug-susceptible to MDR, and from MDR to extensively-drug resistant (XDR). In addition, they found that the katG S315T isoniazid and rpoB S450L rifampicin resistance mutations were dominant across the different MTBC lineages. CONCLUSIONS: Our molecular epidemiological analyses identified the spread of MDR-TB to Ireland from other jurisdictions and its potential to evolve to XDR-TB.

Identification of emergent bla CMY-2 -carrying Proteus mirabilis lineages by whole-genome sequencing.

Whole-genome sequencing of 24 Proteus mirabilis isolates revealed the clonal expansion of two cefoxitin-resistant strains among patients with community-onset infection. These strains harboured bla CMY-2 within a chromosomally located integrative and conjugative element and exhibited multidrug resistance phenotypes. A predominant strain, identified in 18 patients, also harboured the PGI-1 genomic island and associated resistance genes, accounting for its broader antibiotic resistance profile. The identification of these novel multidrug-resistant strains among community-onset infections suggests that they are endemic to this region and represent emergent P. mirabilis lineages of clinical significance.

Molecular epidemiology of Mycobacterium abscessus complex isolates in Ireland.

BACKGROUND: The Mycobacterium abscessus complex are the rapidly growing mycobacteria (RGM) most commonly causing lung disease, especially in cystic fibrosis (CF) patients. Ireland has the world's highest CF incidence. The molecular epidemiology of M. abscessus complex in Ireland is unreported. METHODS: We performed rpoB gene sequencing and multi-locus sequence typing (MLST) on M. abscessus complex strains isolated from thirty-six patients in 2006-2012 (eighteen known CF patients). RESULTS: Twenty-eight strains (78%) were M. abscessus subsp. abscessus, eight M. abscessus subsp. massiliense, none were M. abscessus subsp. bolletii. Sequence type 1 (ST1) and ST26 (M. abscessus subsp. abscessus) were commonest. Seven M. abscessus subsp. abscessus STs (25%) were novel (two with novel alleles). Seven M. abscessus subsp. massiliense STs were previously reported (88%), including two ST23, the globally successful clone. In 2012, of 552 CF patients screened, eleven were infected with M. abscessus complex strains (2%). CONCLUSIONS: The most prevalent M. abscessus subsp. abscessus and M. abscessus subsp. massiliense strains in Ireland belong to widely-distributed STs, but there is evidence of high M. abscessus subsp. abscessus diversity.

Whole-genome sequencing improves discrimination of relapse from reinfection and identifies transmission events among patients with recurrent Clostridium difficile infections.

BACKGROUND: Recurrent Clostridium difficile infection (CDI) represents a significant healthcare challenge. Patients may suffer multiple episodes of CDI with the index strain (relapse) or become infected by another strain acquired nosocomially (reinfection). AIM: We aimed to characterize C. difficile isolates causing recurrent CDI at a tertiary referral hospital by whole-genome sequencing (WGS) to assess strain similarities at the highest level of genetic resolution and accurately detect relapse, reinfection, and putative strain transmission events. METHODS: An 18-month prospective study of recurrent CDI was undertaken. Clostridium difficile was cultured from stool samples collected longitudinally from any patients suffering ≥2 clinically defined CDI episodes. Patient demographics and clinical data were recorded, and strain relatedness investigated by both polymerase chain reaction (PCR)-based ribotyping and WGS. FINDINGS: Nineteen patients were identified with ≥2 clinically defined CDI episodes who cumulatively suffered 39 recurring CDI episodes (58 total episodes). Patients had a median length of stay (LOS) of 144 days and experienced between two and seven CDI episodes. Ribotyping indicated 27 apparent same-strain relapses, five reinfections and the predominance of ribotypes 078 (ST-11) and 020 (ST-2). WGS allowed characterization of relapse with increased certainty and identified emergent within-strain single nucleotide variants (SNVs) with potential functional impact on diverse genes. Shared ribotypes among 14 patients with recurrent CDI suggested 10 possible patient-to-patient transmission events. However, WGS revealed greater diversity at the sub-ribotype level, excluding all but four transmission events. CONCLUSION: WGS exhibits several advantages over PCR-based ribotyping in terms of its ability to distinguish relapse from reinfection, to identify patient-to-patient transmission events, and to exact fine structure characterization of recurrent CDI epidemiology. This offers the potential for more focused infection prevention strategies to eliminate strain transmission among patients with recurrent CDI.

Reexpansion hypotension. A complication of rapid evacuation of prolonged pneumothorax.

Three cases of hypotension are described that followed rapid evacuation of persistent unilateral pneumothorax. Common features included the presence of a pneumothorax for approximately one week before treatment commenced and profuse unilateral reexpansion edema, a rising hematocrit reading, hypotension, and anuria after evacuation of the pneumothorax in spite of a relatively normal pulmonary capillary wedge pressure. In one case, cardiac output was measured and found to be low (1.54 and 1.65 L/min/sq m), with a pulmonary capillary wedge pressure of 10 to 14 mm Hg. Death due to cardiovascular collapse occurred in one patient; ischemic colitis, acute renal failure, disseminated intravascular coagulation, and ischemic necrosis of both humeral heads occurred in another. The cases presented and the literature reviewed suggest that cardiovascular compromise was the end result of the combined effects of intravascular volume depletion and myocardial depression.

Transmission of toxoplasmosis by bone marrow transplant associated with Campath-1G.

A 12-year-old boy in third remission acute lymphoblastic leukaemia was given a mismatched transplant from his mother. He suffered prolonged neutropenia and pyrexia which was only finally diagnosed as toxoplasmosis using molecular biology methods and by his response to appropriate treatment. This was probably transmitted by bone marrow transplant since maternal immune T cells were removed by the use of Campath-1G and treatment with cyclosporin A probably prevented his IgM immune response and impeded the diagnosis.

Increased serum IgE concentrations during infection and graft versus host disease after bone marrow transplantation.

Serum IgE concentrations estimated in 25 bone marrow transplant recipients during episodes of infection or graft versus host disease, or both, were raised not only in some patients with acute graft versus host disease but also in many patients with infection. Raised values were not seen in chronic graft versus host disease. The routine estimation of serum IgE in bone marrow transplant recipients had minimal value because of the lack of specificity of the IgE response.

Value of serum C-reactive protein measurement in the management of bone marrow transplant recipients. Part I: Early transplant period.

Serum C-reactive protein concentrations were measured serially during the early transplant period in 68 bone marrow recipients transplanted for leukaemia (34), chronic granulocytic leukaemia (2), severe aplastic anaemia (6), and various inborn errors of metabolism (26). There were 116 clearly documented episodes of infection or acute graft versus host disease or both. Serum C-reactive protein concentrations in patients with viral (11) or fungal infection (6) were normal or only slightly raised. In 32 patients with isolated acute graft versus host disease, only three (10%) showed serum C-reactive protein concentrations above 40 mg/l. Values greater than 40 mg/l were strongly suggestive of bacterial infections and values above 100 mg/l were seen only in patients (43) with bacterial infections with or without acute graft versus host disease. These findings suggest that serum C-reactive protein concentrations are valuable both for diagnosis and monitoring of such infections.

Value of serum C-reactive protein measurement in the management of bone marrow transplant recipients. Part II: Late post-transplant period.

Seventeen bone marrow recipients transplanted for acute leukaemia (8), chronic leukaemia (1), severe aplastic anaemia (3), and various inborn errors of metabolism (5) had 22 episodes of documented infection in the late (greater than 3 months) post-transplant period. Serum C-reactive protein concentrations were considerably increased in patients with bacterial infections, but not in those with viral or fungal infections. Serum C-reactive protein values were normal in 20 patients transplanted for acute leukaemia (12), chronic leukaemia (1), severe aplastic anaemia (2), and various inborn errors of metabolism (5) who had active chronic graft versus host disease but no evidence of infection. These findings indicate that serum C-reactive protein concentrations are useful in the diagnosis and monitoring of bacterial infections even in the presence of chronic graft versus host disease.

The inhibitory effect of serum on the growth of Torulopsis glabrata.

Normal human plasma and serum were found to inhibit the growth of Torulopsis glabrata and, to a lesser extent, other yeasts. The factor responsible for the inhibition of T. glabrata was not dialysable, was heat stable at 56 degrees C for up to 4 h and could be partly removed by absorption with viable T. glabrata but not Candida albicans. It was fungistatic at low concentrations and fungicidal at high concentrations, stable up to 4 years between -20 degrees C and -70 degrees C, but for only a few weeks at 4 degrees C. Studies with Cohn fractions of serum showed that the inhibitory components were in either the alpha or beta globulin fraction or both. The combined effects of transferrin and IgM accounted for about 70% of the total inhibition observed. We were unable to identify the component responsible for the residual inhibition of growth. The inhibitory effect was totally neutralised by tetracyclines, quinolones, sulphamethoxazole and by very low concentrations of polyenes, imidazoles and 5-fluorocytosine.

Detection of antibodies to common antigens of pathogenic and commensal Neisseria species.

Sera from 29 children and six adults were used to investigate the nature of antigenic cross-reactivity between Neisseria polysaccharea, N. lactamica and N. meningitidis B,15P1.16 by immunoblotting. Major common antigens of 68-70 Kda, 60-65 Kda and 15-20 Kda were detected. Antibody directed against them uniformly decreased after absorption of the sera with the three different Neisseria species. Antigens of 55 Kda and 35 Kda specific to N. meningitidis, and one of 43 Kda specific to N. lactamica, were also demonstrated. Antibody against all antigens was more prevalent in bactericidal than in non-bactericidal sera, although these differences were statistically not significant. Differences in antibody prevalence between carriers of Neisseria spp. and non-carriers of these organisms were even less marked. Examination of sera by whole-cell enzyme-linked immunosorbent assay against N. meningitidis B,15P1.16 and N. lactamica gave an absorbance ratio of 1:1. Only four sera from children showed no reactivity against the meningococcal strain. These common antigens are likely to be important in vaccine development.

The phenotypic relationship of Neisseria polysaccharea to commensal and pathogenic Neisseria spp.

Eight of 22 non-capsulate strains of Neisseria meningitidis previously isolated from primary school children were re-identified as N. polysaccharea by aminopeptidase reactions and polysaccharide production. N. polysaccharea was not identified amongst 91 non-capsulate strains of N. meningitidis isolated from adults attending the Genito-urinary Medicine clinic, Westminster Hospital, London. The biochemical reactions of N. polysaccharea strains were similar to those of N. lactamica and N. gonorrhoeae, but N. polysaccharea could be distinguished from these organisms by examination of beta-galactosidase activity, carbohydrate reactions and polysaccharide production. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis revealed closer similarity of N. polysaccharea to N. lactamica than to the pathogenic Neisseria spp. An additional finding was variation in the position of one of the major proteins of N. lactamica in the 34-39-Kda region.

Disseminated Aspergillus terreus infection arising from cutaneous inoculation treated with caspofungin.

A case of disseminated Aspergillus terreus infection in a patient with prolonged neutropenia after stem cell transplant for myeloma is reported. The isolate was resistant to amphotericin B in vitro, and the patient was successfully managed with surgical debridement and the recently licensed antifungal agent caspofungin. There are many challenges associated with treating invasive aspergillosis, particularly that due to A. terreus, and the early use of caspofungin should be considered.