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Infant high-grade gliomas appear clinically distinct from their counterparts in older children, indicating that histopathologic grading may not accurately reflect the biology of these tumors. We have collected 241 cases under 4 years of age, and carried out histologic review, methylation profiling, and custom panel, genome, or exome sequencing. After excluding tumors representing other established entities or subgroups, we identified 130 cases to be part of an “intrinsic” spectrum of disease specific to the infant population. These included those with targetable MAPK alterations, and a large proportion of remaining cases harboring gene fusions targeting ALK (n = 31), NTRK1/2/3 (n = 21), ROS1 (n = 9), and MET (n = 4) as their driving alterations, with evidence of efficacy of targeted agents in the clinic. These data strongly support the concept that infant gliomas require a change in diagnostic practice and management.Significance:Infant high-grade gliomas in the cerebral hemispheres comprise novel subgroups, with a prevalence of ALK, NTRK1/2/3, ROS1, or MET gene fusions. Kinase fusion–positive tumors have better outcome and respond to targeted therapy clinically. Other subgroups have poor outcome, with fusion-negative cases possibly representing an epigenetically driven pluripotent stem cell phenotype.See related video: https://vimeo.com/438254885See related commentary by Szulzewsky and Cimino, p. 904.This article is highlighted in the In This Issue feature, p. 890

IntroductionAn emerging pre-clinical approach for the treatment of pharmacoresistant epilepsy is targeting the microRNA (miRNA) system. MiRNAs are short noncoding RNAs that suppress gene expression at the post-transcriptional level. Targeting miRNAs, which is possible using antisense oligonucleotide ‘antimiRs’ can produce broad effects on gene expression suited to the complex pathophysiology in temporal lobe epilepsy. Potent anti-seizure and disease- modifying effects have been reported for antimiRs targeting microRNA-134 (antimiR-134). To date, however, pre-clinical testing has been performed using in vitro cell cultures and rodent models. It is uncertain how well this approach will translate to the clinic. Here, we develop an antimiR testing platform in human brain tissue sections.MethodologyHuman brain specimens were obtained with consent from patients undergoing resective surgery to treat focal drug-resistant epilepsy. Neocortical specimens were submerged in modified artificial cerebrospinal fluid (ACSF), dissected for clinical neuropathological examination, and unused material transferred for sectioning. Individual tissue sections were incubated in oxygenated ACSF, containing either antimiR-134 or a non-targeting control antimiR, for 24 hours at room temperature. RNA integrity was assessed using BioAnalyzer processing, and individual miRNA levels measured using RT-qPCR.ResultsACSF transport had no obvious impact on any clinical neurosurgical or neuropathological procedure and specimens were confirmed to be viable following this process. RNA was well- preserved by transportation of specimens in ACSF, with RNA integrity scores significantly higher than tissue transported without ACSF. AntimiR-134 mediated a specific and dose- dependent knockdown of miR-134 in human neocortical sections, with approximately 75% reduction of miR-134 at 1 µM and 90% reduction at 3 µM. These doses did not have off- target effects on expression of a selection of three other miRNAs (miR-10, miR-129 or miR- 132).SignificanceThis is the first demonstration of antimiR-134 effects in live human brain tissues. The findings lend further support to the preclinical development of miR-134 and offer a flexible platform for the pre-clinical testing of antimiRs, and other antisense oligonucleotide therapeutics, in human brain.Key pointsASO antimiRs are promising treatments for pharmacoresistant epilepsyWe developed a pipeline to preserve live human neocortical brain specimens from people undergoing resective surgeryRNA integrity was sufficient to measure miRNA levels in human brain tissues transported in modified ACSFIncubation of acute human neocortical specimens in antimiR-134 resulted in potent and specific reduction in miR-134 levelsAcute human brain slices are a promising model for testing ASOs

The Neuropathology of Human Parechovirus (HPeV) is not widely described due to the relatively recent discovery of the virus combined with a limited number of autopsy case reports. We report the case of an infant boy born at 38 weeks who, six days after birth, presented with fever and severe neurological dysfunction. Human Parechovirus Type 3 (HPeV3) RNA was detected in his cerebrospinal fluid (CSF) and blood. He died five days after his initial presentation. Neuropathologic examination demonstrated multicystic encephalomalacia (ME). This case report confirms that white matter pathology is dominant in HPeV3 infection. A unique feature, of HPeV encephalomalacia is absence of CSF pleocytosis and minimal inflammation in the meninges. The findings permit comment on the pathogenesis of brain injury by this virus.

Summary Background Ireland has been shown to have the highest rate of vancomycin-resistant enterococci (VRE) in cases of bacteraemia in Europe, according to a report in 2014 from the European Antimicrobial Resistance Surveillance System Network. Aim To investigate the prevalence of VRE gut colonization in a cohort of patients in 2014 at Cork University Hospital (CUH) by performing a cross-sectional study using faecal samples submitted to the microbiology laboratory for routine investigation from both hospital inpatients and community-based patients. Methods Faeces were examined for VRE colonization using selective cultivation, antimicrobial susceptibility testing, and speciation using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. All VRE isolates were evaluated by molecular means for resistance determinants, type, and Insertion Sequence 16 as an indicator of Clonal Complex 17 (CC17). Findings From the 350 specimens investigated, 67 (19.1%) specimens were positive for VRE [95% confidence interval (CI): 15.0–23.2]. The prevalence of VRE colonization among CUH patients tested in this study ( N = 194) was 31.4% (95% CI: 24.7–38.1). By contrast, the general practitioner patient samples ( N =29) showed a prevalence of 0%, whereas 22.2% of samples from other hospitals ( N =27) were positive for VRE. All isolates were Enterococcus faecium (VRE fm ) and were indicated to contain CC17, though with considerable heterogeneity among the isolates. Conclusion This high prevalence goes some way towards providing an explanation for the current high rates of VRE bacteraemia in Ireland, as well as highlighting the benefits of screening and enhanced infection control practices by all hospitals to control the high rates of VRE observed.

We appreciate the opportunity to respond to the letter by Ackerman et al. [1], andwe greatly appreciate the effort of the authors of the letter to communicate their concerns to us. We attempt to address these concerns below. Please note that Ms. Laura Crandall has recused herself from this response due to a conflict of interest as a member of the SUDC Foundation, since the letter writers represent the board of the foundation. 1. The major concerns raised in the letter are that the design of the study was not population-based, the cases were accrued retrospectively, and there was inconsistency across medical examiners on the information available to us for review. We have organized our response to address these major and related concerns together first. In the response, we summarize the critique of the authors in italics and then reiterate (in quotation marks) and clarify the text of our original paper in which the points questioned were addressed.

Sudden unexplained death in childhood (SUDC), while rare, accounts for an important fraction of unexpected deaths in children >1 year of age. Previously we reported an association between febrile seizures, hippocampal maldevelopment, and sudden, unexpected deaths in young children (1–6 years), termed “hippocampal maldevelopment associated with sudden death (HMASD).” Here, we characterize in greater detail the hippocampal pathology in a large cohort of cases (n = 42) of this entity, and attempt to define possible new entities responsible for sudden, unexplained death in young children without HMASD/febrile seizure phenotypes. We performed comparative analysis on cases, which we classified in a cohort of 89 sudden and unexpected deaths as HMASD, explained deaths, SUDC with febrile seizure phenotype (SUDC-FS) but without hippocampal pathology, and SUDC (without hippocampal pathology or febrile seizure phenotype). The frequency of each subgroup was: HMASD 48 % (40/83); SUDC 27 % (22/83); SUDC-FS 18 % (15/83); explained 7 % (6/83). HMASD was characterized clinically by sudden, sleep-related death, term birth, and discovery in the prone position. Key morphologic features of HMASD were focal granule cell bilamination of the dentate gyrus with or without asymmetry and/or malrotation of the hippocampus, associated with significantly increased frequencies of 11 other developmental abnormalities. We identified no other distinct phenotype in the unexplained categories, except for an association of febrile seizures without hippocampal maldevelopment. HMASD is a distinct clinicopathologic entity characterized by a likely developmental failure of neuronal migration in the dentate gyrus. Future research is needed to determine the causal role of HMASD in sudden death in early childhood.

Purpose The purpose of this study was to determine the major subcategories and clinicopathologic features of sudden unexpected death in young children in a large retrospective cohort, and to confirm the association of sudden unexplained death in children (abbreviated by us for unexplained deaths as SUDC) with hippocampal pathology and/or febrile seizures. Methods We undertook analysis of a retrospective cohort of 151 cases, of which 80 % (121/151) were subclassified as SUDC, 11 % (16/151) as explained, 7 % (10/151) as undetermined, and 3 % (4/151) as seizure-related. Results There were no significant differences between SUDC and explained cases in postnatal, gestational, or postconceptional age, frequency of preterm birth, gender, race, or organ weights. In contrast, 96.7 % (117/121) of the SUDC group were discovered during a sleep period compared to 53.3 % (8/15) of the explained group (p

SUMMARYIn October 2012, an outbreak of gentamicin-resistant, ciprofloxacin non-susceptible extended-spectrumβ-lactamase (ESBL)-producingKlebsiella pneumoniaeoccurred in a neonatal intensive care unit in Ireland. In order to determine whether the outbreak strain was more widely dispersed in the country, 137 isolates ofK. pneumoniaewith this resistance phenotype collected from 17 hospitals throughout Ireland between January 2011 and July 2013 were examined. ESBL production was confirmed phenotypically and all isolates were screened for susceptibility to 19 antimicrobial agents and for the presence of genes encodingblaTEM,blaSHV,blaOXA, andblaCTX-M; 22 isolates were also screened forblaKPC,blaNDM,blaVIM,blaIMPandblaOXA-48genes. All isolates harbouredblaSHVandblaCTX-Mand were resistant to ciprofloxacin, gentamicin, nalidixic acid, amoxicillin-clavulanate, and cefpodoxime; 15 were resistant to ertapenem, seven to meropenem and five isolates were confirmed as carbapenemase producers. Pulsed-field gel electrophoresis of all isolates identified 16 major clusters, with two clusters comprising 61% of the entire collection. Multilocus sequence typing of a subset of these isolates identified a novel type, ST1236, a single locus variant of ST48. Data suggest that two major clonal groups, ST1236/ST48 (CG43) and ST15/ST14 (CG15) have been circulating in Ireland since at least January 2011.

Sudden infant death syndrome (SIDS) and sudden unexplained death in childhood (SUDC) are defined as sudden death in a child remaining unexplained despite autopsy and death scene investigation. They are distinguished from each other by age criteria, i.e. with SIDS under 1 year and SUDC over 1 year. Our separate studies of SIDS and SUDC provide evidence of shared hippocampal abnormalities, specifically focal dentate bilamination, a lesion classically associated with temporal lobe epilepsy, across the 2 groups. In this study, we characterized the clinicopathologic features in a retrospective case series of 32 children with sudden death and hippocampal formation (HF) maldevelopment. The greatest frequency of deaths was between 3 weeks and 3 years (81%, 26/32). Dentate anomalies were found across the pediatric age spectrum, supporting a common vulnerability that defies the 1-year age cutoff between SIDS and SUDC. Twelve cases (38%) had seizures, including 7 only with febrile seizures. Subicular anomalies were found in cases over 1 year of age and were associated with increased risk of febrile seizures. Sudden death associated with HF maldevelopment reflects a complex interaction of intrinsic and extrinsic factors that lead to death at different pediatric ages, and may be analogous to sudden unexplained death in epilepsy.

Enteric adenoviruses have been shown to be a substantial cause of pediatric gastroenteritis in various parts of the world, and are considered to be the second most common cause of viral gastroenteritis, next to rotavirus in young children. Genetic characterization of 95 adenovirus isolates obtained from patients with acute gastroenteritis between 2002 and 2007 from the southern regions of Ireland, were characterized by PCR analysis, restriction endonuclease (RE) analysis and sequencing analysis. All isolates were found to be of adenovirus type 41 origin. Genetic analysis of seven hypervariable regions (HVRs) located within the hexon gene has revealed a high level of amino acid sequence homology in samples over the course of this study, with a very close relationship to the D22 genome type. The D22 genome type has been detected in several other countries, thus suggesting Irish isolates have common genome types with other stains worldwide. This is the first such study undertaken in the south of Ireland, to type and genetically characterize adenoviral gastroenteritis isolates, and has revealed a high level of conservation within the isolated analyzed.

There were two parts to this study. Part 1 evaluated the epidemiology of Candida bloodstream isolates within the Southern Health Board (SHB) of Ireland from 1992 to 2003 by retrospective surveillance of all such isolates of patients reported from SHB hospitals to our laboratory database during that period. Part 2 reviewed candidaemia cases occurring in Cork University Hospital (CUH) from 1999 to 2003 using surveillance of all positive blood culture isolates in CUH microbiology laboratory during the 5-year period. In part 1, 250 Candida bloodstream isolates were reported in the SHB over 12 years. There was a pattern of decreasing percentage of C. albicans with time. Whereas in part 2, 63 cases of candidaemia were identified in CUH from 1999 to 2003. Candida albicans constituted 50% of all isolates, while C. parapsilosis and C. glabrata accounted for 21.2% and 18.2% respectively. Average annual incidence rate was 0.48 episodes/1000 admissions and 0.70 episodes/10 000 patient-days. Vascular catheters were the commonest source of candidaemia (61.9%) followed by the urinary tract (12.7%). Risk factors included exposure to multiple antibiotics (75%); central vascular catheterization (73%); multiple colonization sites (71%); severe gastrointestinal (GI) dysfunction (54%) and acute renal failure (43%). Crude 7-day and 30-day mortality rates were 20.6% and 39.7% respectively. Logistic regression multivariate analysis identified the following to be independent predictors for mortality: age > or =65 years [odds ratio (OR) 7.2, P = 0.013]; severe GI dysfunction (OR 10.6, P = 0.01); acute renal failure (OR 7.6, P = 0.022); recent/concurrent bacteraemia (OR 5.2, P = 0.042); endotracheal intubation (OR 7.7, P = 0.014); while major surgery was associated with a better prognosis (OR 0.05, P = 0.002). Appropriate antifungal treatment was also found to be associated with survival (Fisher's exact test, P < 0.001). The epidemiology of Candida bloodstream isolates within our health board had changed over the years. Incidence and mortality of candidaemia were relatively high in our hospital. Dysfunction of major organ systems and recent bacteraemia were found to predict mortality.

Objectives: In this study a large random collection (n = 378) of Irish thermophilic Campylobacter isolates were investigated for the presence of integrons, genetic elements associated with the dissemination of anti- microbial resistance. Methods: Purified genomic DNA from each isolate was analysed by PCR for the presence of class 1 inte- grons. Four gene cassette-associated amplicons were completely characterized. Results: Sixty-two of the isolates possessed a complete class 1 integron with a recombined gene cassette located within a 1.0 kb amplicon containing an aadA2 gene. This cassette was present in both Campylo- bacter jejuni and Campylobacter coli isolates and following sequence analysis was shown to be similar to sequences recently reported in Salmonella enterica Hadar and on an 85 kb plasmid conferring quinolone resistance in Escherichia coli. Conclusions: Aminoglycoside aadA2-encoding class 1 integrons were identified among unrelated Campy- lobacter spp. Amino acid sequence comparisons revealed identical structures in both Salmonella and E. coli. The presence of class 1 integrons in Campylobacter spp. may be significant should these organisms enter the food chain and especially when antimicrobial treatment for severe infections is being considered.

Burkholderia cepacia is a Gram-negative bacterium associated with increasing morbidity and mortality and is readily transmitted among infected cystic fibrosis (CF) patients. The B. cepacia complex consists of five distinct subgroups, termed genomovars. A collection of 17 presumptive B. cepacia isolates, obtained from three national CF referral centres located in different geographical regions in Ireland, was studied. The aim of this study was to investigate these isolates using molecular subtyping protocols for evidence of genetic relationships and for the presence of antibiotic resistance-encoding class 1 integron structures.Genomovar classifications were assigned to each isolate based on HaeIII enzyme profiles of their recA locus. Genetic relationships among this collection were also assessed after restriction fragment length polymorphism (RFLP)-mediated analysis of the 16S rDNA locus and DNA amplification fingerprinting (DAF). The surface expression of the cable pilus gene (cblA) may facilitate an early step in the infection process. All isolates were tested by amplification strategies for this marker. Burkholderia cepacia is known to be resistant to several antimicrobial agents. Resistance typing showed that the majority were resistant to three or more common antimicrobial agents. Five of the 17 isolates were resistant to sulphonamide, a characteristic linked with the presence of class 1 integrons. Gene cassettes containing beta-lactamase (oxa) and aminoglycoside acetyltransferase (aac(6')-1a) encoding genes were identified by polymerase chain reaction amplification.Most of the isolates in this study were classified as genomovar III and were indistinguishable based on their corresponding 16S rDNA-RFLP profiles, whilst DAF further subtyped the collection. The cblA marker was identified in 47% of the isolates, many of which clustered in the genomovar III group. Class 1 integrons with recombined gene cassettes containing bla-OXA and aac(6')-1a genes were identified.This study demonstrates the application of molecular methods to investigate B. cepacia, a well-recognized human pathogen, cultured from Irish CF patients. Genomovar III was the most common genomic type identified. DNA fingerprinting further subtyped the latter isolates, facilitating a more detailed description of the molecular epidemiology. Drug resistance in these organisms can be explained, at least in part, by the presence of class 1 integrons. Development of targeted infection control strategies could be facilitated using these applied methods.

Background. We estimated the disease burden caused by rotavirus hospitalizations in the Republic of Ireland by using national data on the number of hospitalizations for diarrhea in children and laboratory surveillance of confirmed rotavirus detections. Methods. We examined trends in diarrheal hospitalizations among children

Antimicrobial resistance (R) typing and DNA Amplification Fingerprinting (DAF) of a random collection of 84 Irish thermophilic Campylobacter isolates is described. The collection included human, veterinary (porcine) and poultry isolates cultured between 1996 and 1998 in the Cork region of Ireland. Biochemical and molecular methods were used to identify Campylobacter jejuni and Camp. coli. Many of these isolates were simultaneously resistant to several common antimicrobial agents. In particular, resistance to ampicillin, spectinomycin, sulphafurazole and tetracycline was common. A total of 74 DAF profiles was identified among the study collection, showing a high degree of diversity. Dendrogram analysis of the DNA patterns identified three main clusters at the 50% similarity level, which included two clusters of Camp. coli and a third containing a mixture of Camp. jejuni and Camp. coli.

Veillonella species is a rare cause of endocarditis. We report a case of a 49-year-old man with Veillonella parvula prosthetic valve endocarditis who presented with acute cardiac failure due to valvular dehiscence. His clinical course was complicated by cortical blindness and limb paresis as a result of cerebral embolism. The endocarditis was successfully treated with urgent valve replacement surgery and a prolonged course of metronidazole.

Norovirus (NoV) gastroenteritis occurs in all age groups and is the most common cause of gastroenteritis in the community. However, detection methods and rates vary widely, and few data are available to compare these, particularly in Ireland. Detection of noroviruses through antigen and molecular-based strategies was carried out on 135 suspected NoV-positive samples, collected over the course of three NoV outbreaks, from 2002 to 2006, in the southern region of Ireland. A commercially available ELISA and a panel of six primer sets were evaluated to determine their suitability for NoV detection in Irish clinical samples. The key findings of this study were the detection of both GGI and GGII noroviruses by ELISA, but the detection of only GGII noroviruses by RT-PCR. In addition to this, a variation in the levels of detection from 9.4 % to 17.3 % was observed for conventional PCR assays, while a detection rate of 46.3 % was observed for the real-time PCR assay. A proportion (17.8 %) of samples were found to be negative by all detection strategies, suggesting the possibility of reporting false positives for these samples or low-copy positives that do not often repeat. Sequencing information from selected samples also revealed nucleotide polymorphisms, compromising efficient primer binding in the case of one primer pairing. Phylogenetic analysis of the partial polymerase gene identified NoV GII.4 as the dominant genotype, in accordance with previous NoV studies in Ireland. Investigating the NoV diversity of the circulating strains and the dynamics of strain replacement is important to better assess the efficacy of future NoV vaccines and to facilitate the early detection of changes in circulating NoV strains.

Objective: To compare the incidence of tuberculosis due to Mycobacterium bovis in humans to the prevalence of M. bovis infection in cattle in south-west Ireland and discuss possible links between them. Setting: In the south-west region of Ireland, a mixed urban and rural community (pop. 536 000), there is a residuum of human tuberculosis caused by M. bovis . Methods: A retrospective analysis of the incidence of culture-positive M. bovis disease in humans in south-west Ireland from 1983 to 1994 and of the results of tuberculin testing in cattle from 1978 to 1994 for the same region. Results: One to five cases of human tuberculosis due to M. bovis were recorded per year while the overall prevalence of bovine infection fell gradually during the period of study from 467 tuberculin-positive animals per 100 000 cattle tested in 1983 to 158 per 100 000 in 1994. Conclusion: The low incidence plateau of human tuberculosis due to M. bovis together with the decline in prevalence of animal infection in the overall period studied suggest a cut-off in the animal to human chain of infection at two points; the animal source and the ingestion of (now pasteurized) milk. This would suggest that disease in humans is now due to reactivation of previous foci of infection which were acquired when milk pasteurization was not compulsory. Based on this, we would anticipate a further reduction and possible elimination of human tuberculosis due to M. bovis in this region in the next 10–15 years.

Varicella zoster virus (VZV) is an occupational hazard for a percentage of health care staff. Nine hundred and seventy staff members attending the Occupational Health Department at Cork University Hospital took part in the survey. A latex agglutination assay was used to determine the health care workers immune status to VZV. Of the 970 workers tested, 928 (95.7%) were immune to VZV. The sensitivity, specificity and predictive value of an enquiry regarding a history of chicken-pox was determined on a sample of 206 health care workers. The positive predictive value was 95% (119/125) and the negative predictive value was 11% (4/35). The sensitivity of the enquiry was 79% (119/150), the specificity was 40% (4/10), reducing to 61% (119/195) and 36% (4/11) respectively when individuals with uncertain histories were included in the calculations. The advantages and disadvantages of selective staff screening are discussed. In the authors' opinion all health care workers involved in the clinical care of patients should be screened by serology for past VZV infection before taking up duty and those who are susceptible to VZV should be made aware of the risks and health effects associated with VZV if contracted.

Acinetobacter are important emerging nosocomial pathogens. In this paper thirteen Acinetobacter baumanii from intensive care patients were isolated. These were initially typed using the API-20 NE biotyping system and antibiogram analysis. Results obtained using these methods failed to convincingly characterise the organisms. In this report a method to characterise these Acinetobacter baumanii isolates is described, which utilises a modified polymerase chain reaction (PCR), capable of generating genomic fingerprints, known as DNA amplification fingerprinting (DAF). Purified chromosomal DNA of cultured clinical isolates of Acinetobacter baumanii were subjected to DAF using the M13 universal sequencing primer. Polymorphic DNA bands produced, were visualised after agarose gel electrophoresis and ethidium bromide staining. Results demonstrated that six of the thirteen clinical isolates represented one group and a second group of two isolates displayed identical fingerprint patterns. The remaining four organisms were all unique. This genotype based method is rapid, simple, reproducible and may have potential as a means of specifically typing Acinetobacter spp. allowing the route(s) of nosocomial transmission to be identified and to assess the efficiency of instituted infection control measures.

Acinetobacter species are important nosocomial pathogens. A rapid and sensitive identification system, capable of providing strain identity at the genetic level, is required to identify outbreak strains and facilitate the early implementation of infection control procedures. Repetitive extragenic palindromic (REP) elements, have been identified in numerous bacteria and these genomic sequences provide useful targets for DNA amplification. A method for amplifying inter-REP DNA sequences, REP-multiple arbitrary amplicon profiling (REP-MAAP), is described and applied to 29 Acinetobacter baumannii from clinical samples. Amplified polymorphic DNA patterns were demonstrated for all isolates and those displaying identical REP-MAAP patterns were considered identical at the genetic level. In the spring of 1993, 10 intensive care unit patients had endotracheal colonization with A. baumannii (five with REP-MAAP I and five with REP-MAAP II patterns). These findings suggested nosocomial transmission of organisms which was terminated by standard infection control measures. No further A. baumannii were detected until the winter of 1993 when isolates of different REP-MAAP groups emerged, suggesting that factors other than nosocomial transmission were implicated.

SUMMARYThis study determined the carriage rate and serotype distribution of group BStreptococcus(GBS) in women of child-bearing age in the southern region of Ireland. A total of 2000 vaginal swabs collected in two periods in 2004 and 2006 were examined and revealed a GBS carriage rate of 16·1%. Serotyping of isolates showed that serotypes Ia, II, III, IV, and V were the most prevalent. A high prevalence of serotype IV was found, increasing from 7·6% to 15·2% between 2004 and 2006. Random amplified polymorphic DNA analysis demonstrated considerable genetic heterogeneity in the serotype IV isolates. This serotype should be considered for inclusion in potential vaccines for use in Ireland.

Campylobacter sputorum biovar sputorum is a rarely isolated organism, particularly from human clinical specimens. Its pathogenic potential is unknown. We present here what we believe to be the first report of this organism being isolated from a clinically significant source, an axillary abscess. To our knowledge, this organism has not been reported previously as one of clinical relevance in the U.K.

BACKGROUND—The ligase chain reaction Mycobacterium tuberculosis assay uses ligase chain reaction technology to detect tuberculous DNA sequences in clinical specimens. A study was undertaken to determine its sensitivity and specificity as a primary screening tool for the detection of culture positive tuberculosis. METHODS—The study was conducted on 2420 clinical specimens (sputum, bronchoalveolar lavage fluid, pleural fluid, urine) submitted for primary screening for Mycobacterium tuberculosis to a regional medical microbiology laboratory. Specimens were tested in parallel with smear, ligase chain reaction, and culture. RESULTS—Thirty nine patients had specimens testing positive by the ligase chain reaction assay. Thirty two patients had newly diagnosed tuberculosis, one had a tuberculosis relapse, three had tuberculosis (on antituberculous therapy when tested), and three had healed tuberculosis. In the newly diagnosed group specimens were smear positive in 21 cases (66%), ligase chain reaction positive in 30 cases (94%), and culture positive in 32 cases (100%). Using a positive culture to diagnose active tuberculosis, the ligase chain reaction assay had a sensitivity of 93.9%, a specificity of 99.8%, a positive predictive value of 83.8%, and a negative predictive value of 99.9%. CONCLUSIONS—This study is the largest clinical trial to date to report the efficacy of the ligase chain reaction as a primary screening tool to detect Mycobacterium tuberculosis infection. The authors conclude that ligase chain reaction is a useful primary screening test for tuberculosis, offering speed and discrimination in the early stages of diagnosis and complementing traditional smear and culture techniques.

Objectives : an outbreak of Salmonella tel-el-kebir occurring over a 6-month period is described in this report. This is thefirst outbreak of S. tel-el-kebir in the reported literature. Methods : S. tel-el-kebir was isolated from human faecal samples using conventional laboratory methods. Results : eight patients had S. tel-el-kebir isolated from faeces. All patients were owners of, or in close contact with, pet terrapins. The terrapins were purchased in the same pet shop, where they were imported from America. The epidemiological link with these pets was confirmed, as S. tel-el-kebir was isolated from cloacal swabs from the terrapins, and from terrapin water. Molecular biology studies using DNA amplification fingerprinting (DAF) gave identical fingerprint patterns for all human and terrapin isolates. Conclusions : Salmonellosis associated with exotic pets is a re-emerging disease in the 1990s, and measures to reduce this are discussed.

Serratia marcescens is a frequent nosocomial isolate and is associated with a variety of clinical sources, including blood, urine and sputum, and can cause significant infection. Infections can be difficult to treat due to its resistance to a variety of antimicrobial agents. An investigation of a population of 30 clinical strains revealed the presence of a class 2 integron among nine of the isolates, which represents the first isolation of this integron in Serratia species. This integron contained the gene cassettes dfrA1, sat1 and aadA1, conferring resistance to trimethoprim, streptothricin and streptomycin/ spectinomycin, respectively. One of these isolates also carried a class 1 integron identified by sequence analysis as containing the open reading frames aacC1 (encoding gentamicin resistance), ORFX, ORFY and aadA1. Polymerase chain reaction analysis confirmed the presence of the qac epsilon delta1 and sul1 markers, which are common among class 1 integrons.

A commercial DNA/DNA hybridisation kit for the detection of Escherichia coli heat stable enterotoxin gene sequences was compared to the suckling mouse bioassay using 183 isolates of Escherichia coli from clinical specimens. The gene probe assay had a specificity of 99% and a sensitivity of 90.4% compared to the infant mouse method. Using the colony blot method of preparing the bacterial DNA and a hybridisation temperature of 50 degrees C optimal results were obtained. The gene probe method is not affected by the incubation conditions of the test organisms. It is technically straightforward and can be applied to large numbers of specimens with fewer logistic difficulties than with the bioassay.

Two hundred and thirty fecal specimens were collected from children (up to 5 years of age) admitted with suspected rotaviral gastroenteritis at four Irish hospitals (Cork University Hospital, Mercy Hospital, Cork, Waterford Regional Hospital, and Kerry General Hospital) in the southern region of Ireland, between 2001 and 2004. Following laboratory confirmation of the aetiological agent, the rotavirus G-type was determined in all positive samples by reverse transcriptase-polymerase chain reaction (RT-PCR). The distribution of the G-types (n = 230) over the 3 year period was G1 (31%), G9 (21.8%), G3 (8.7%), G4 (6.5%), and G2 (3.5%). There were many mixed infections which accounted for 28.5% of the collection. G9 emerged as the most prevalent G type (30.1%) in 2001–2002, whilst G3 first emerged in 2002–2003 and accounted for 15.8% of the collection. Notably, G2 strains were present at a very low frequency (3.5%) during 2001–2004, compared to an earlier study (1997–1999), where they accounted for 28.5% of the specimens. A smaller subset of the study collection was similarly P-typed (n = 139). P[8]-type was identified as the most prevalent P-type, accounting for 97.4% (n = 186), while P[4] accounted for just 2.6% (n = 5) of the collection. The low frequency of P[4] coincided with the decrease in G2 strains in circulation. The key finding in this study was the emergence of G3- and G9-serotypes as epidemiologically important rotavirus strains since 1999, and the low prevalence of the previously common G2 strains in Ireland. The profile of rotavirus is changing continuously in Ireland and the implications for a successful vaccination program are discussed. J. Med. Virol. 77:571–578, 2005. © 2005 Wiley-Liss, inc.

We report infection in a 4-year-old girl with cervical adenitis caused by Mycobacteria other than tuberculosis and the subsequent, rapid identification of this isolate as Mycobacterium malmoense , using a combined PCR and DNA probe assay.

Background: Broad-spectrum antibiotics, particularly intravenous cephalosporins, are associated with Clostridium difficile diarrhoea. Diarrhoea due to C. difficile is a growing problem in hospitals, especially among elderly patients. Aim: To establish whether changing an antibiotic policy with the aim of reducing the use of injectable cephalosporins leads to a reduction in the incidence of C. difficile diarrhoea in elderly patients. Design: Retrospective analysis. Methods: A group of patients who were subject to the new antibiotic policy from the period following July 2000, were compared with patients who were admitted prior to July 2000 and were not subject to the new policy. Infections, antibiotic prescriptions and mortality rates were determined from case notes, and C. difficle diarrhoea rates from microbiological data. Results: Intravenous cephalosporin use fell from 210 to 28 defined daily doses ( p < 0.001) following the change in antibiotic policy, with a corresponding increase in piperacillin-tazobactam ( p < 0.001) and moxifloxacin ( p < 0.001) use. The new policy led to a significant reduction in C. difficile diarrhoea cases. The relative risk of developing C. difficile infection with the old policy compared to the new policy was 3.24 (95%CI 1.07–9.84, p = 0.03). Discussion: The antibiotic policy was successfully introduced into an elderly care service. It reduced both intravenous cephalosporin use and C. difficile diarrhoea.

Setting: The Southwest of Ireland (Counties Cork and Kerry) 1987–2000, average population 549 500. Objective: Nontuberculous mycobacteria (NTM) cause significant morbidity worldwide and the study of epidemiology and characteristics helps in their prevention and treatment. This study was performed to determine the incidence of NTM disease in comparison to Mycobacterium tuberculosis ( M. tuberculosis ) and Mycobacterium bovis ( M. bovis ) in Southwest Ireland, over the above time period. Design: A retrospective study was carried out in all human isolates of NTM, M. tuberculosis and M. bovis between 1987 and 2000, in the Southwest Region of Ireland. Results: The mean incidence of NTM (0.4/100 000 population) has risen since 1995, principally of pulmonary Mycobacterium avium intracellulare complex (MAC). The annual incidence of M. tuberculosis in humans over 14 years in the same region was 9.71/100 000 population with a significant reduction since 1994 and M. bovis remained constant at 0.5/100 000 population. Conclusion: The increasing incidence of disease causing NTM noted in Southwest Ireland reflects global data and is surmised to be due to an ageing population, increased incidence related to chronic fibrotic lung disease, and environmental mycobacterial factors.

Measurements were made of the susceptibility to six commonly prescribed antibiotics, including erythromycin, tetracycline and ciprofloxacin, of 130 isolates of Campylobacter jejuni and 15 isolates of Campylobacter coli cultured from human and poultry sources during 2000. The results were compared with the results from a collection of strains isolated between 1996 and 1998. The levels of resistance to erythromycin remained low, 2 per cent and 4.4 per cent for the human and poultry isolates, respectively. Resistance to tetracycline had increased to 31 per cent and 24.4 per cent from 13.9 per cent and 18.8 per cent for the human and poultry isolates, respectively. However, the resistance to ciprofloxacin of the strains isolated during 2000 had increased to 30 per cent, whereas between 1996 and 1998 there had been no resistance to this agent among human isolates, and only 3-1 per cent resistance among poultry isolates. The molecular basis for this resistance has been shown to be the result of a single amino acid substitution, Thr-86-lle, in the gyrA subunit of DNA gyrase in C jejuni . A subset of 59 isolates was tested by molecular methods and all of the 25 phenotypically resistant isolates possessed this substitution. None of the human isolates had been treated with ciprofloxacin before their laboratory isolation.

A restriction fragment length polymorphism (RFLP) detection assay was developed to examine the genetic relationship(s) among VP7-encoding genes from 100 Irish rotavirus isolates and 30 randomly selected global rotavirus isolates (from the current databases). RFLP analysis of the VP7 gene segments was performed independently with three enzymes ( Rsa I, Alu I, and Eco RV) in separate reactions by direct digestion of the DNA product amplified by reverse transcriptase (RT)-mediated PCR (RT-PCR) or by using computational methods. Thirty-six RFLP patterns were identified for all 130 strains, and of these, only nine patterns were associated with the Irish isolates. A correlation between the G type of the Irish isolates and certain single or combined enzyme profiles was apparent. These data suggested that the Irish wild-type rotavirus population was homogeneous and could be distinguished by RFLP analysis from global isolates of the same serotype(s). The deduced amino acid sequences of the VP7 RT-PCR products from six Irish isolates known to be of the G serotype revealed significant amino acid substitutions within major antigenic regions. In addition, these data identified the existence of at least two genetic lineages within serotype G1 strains which were distinguishable by RFLP analysis.

A collection of three hundred thirty rotavirus-positive stool samples from children with diarrhea in the southern and eastern regions of Ireland between 1997 and 1999 were submitted to the Molecular Diagnostics Unit of the Cork Institute of Technology, Cork, Ireland, for investigation. These strains were characterized by several methods, including polyacrylamide gel electropherotyping and G and P genotyping. A subset of the G types was confirmed by nucleic acid sequencing. The most prevalent types found in this collection included G1P[8] (n = 106; 32.1%), G2P[4] (n = 94; 28.5%), and G4P[8] (n = 37; 11.2%). Novel strains were also detected, including G1P[4] (n = 19; 5.8%), and G4P[4] (n = 2; 0.6%). Interestingly, mixed infections accounted for 18.8% (n = 62) of the total collection, with only 3% (n = 10) which were not G and/or P typeable. Significantly, six G8 and five G9 strains were identified as part of mixed infections. These strains have not previously been identified in Irish children, suggesting a greater diversity in rotavirus strains currently circulating in Ireland.

Salmonella enterica is among the principal etiological agents of food-borne illness in humans. Increasing antimicrobial resistance in S. enterica is a cause for worldwide concern. There is concern at present in relation to the increasing incidence of human infection with antimicrobial agent-resistant strains of S. enterica serotype Typhimurium, in particular of phage type DT104. Integrons appear to play an important role in the dissemination of antimicrobial resistance genes in many Enterobacteriaceae including S. enterica . In this study the antimicrobial susceptibilities and phage types of 74 randomly collected strains of S. enterica serotype Typhimurium from the Cork region of southern Ireland, obtained from human, animal (clinical), and food sources, were determined. Each strain was examined for integrons and typed by DNA amplification fingerprinting (DAF). Phage type DT104 predominated ( n = 48). Phage types DT104b ( n = 3), -193 ( n = 9), -195 ( n = 6), -208 ( n = 3), -204a ( n = 2), PT U302 ( n = 1), and two nontypeable strains accounted for the remainder. All S. enterica serotype Typhimurium DT104 strains were resistant to ampicillin, chloramphenicol, streptomycin, Sulfonamide Duplex, and tetracycline, and one strain was additionally resistant to trimethoprim. All DT104 strains but one were of a uniform DAF type (designated DAF-I) and showed a uniform pattern of integrons (designated IP-I). The DT104b and PT U302 strains also exhibited the same resistance phenotype, and both had the DAF-I and IP-I patterns. The DAF-I pattern was also observed in a single DT193 strain in which no integrons were detectable. Greater diversity of antibiograms and DAF and IP patterns among non-DT104 phage types was observed. These data indicate a remarkable degree of homogeneity at a molecular level among contemporary isolates of S. enterica serotype Typhimurium DT104 from animal, human, and food sources in this region.

Editor,—Ocular toxoplasmosis is the most frequent infectious cause of chorioretinal inflammation in immunocompetent individuals.1 Diagnosis is usually made by observing the typical fundus lesion, by detecting the presence of anti- Toxoplasma antibodies in the serum, and by excluding other causes of necrotising fundus lesions.2In unusual cases, invasive procedures may be required to aid diagnosis.3 ### CASE REPORT A 17 year old white male presented complaining of floaters and reduced visual acuity in the left eye. Visual acuity was 6/9 in the left eye, 6/6 in the right. Examination revealed moderate anterior chamber activity, marked vitritis, and an active retinochoroiditis adjacent to an area of old chorioretinal scarring inferonasal to the optic disc. A diagnosis of ocular toxoplasmosis was suspected, and topical and oral steroids, …