Genetic relationships between Candida albicans strains isolated from dental plaque, trachea, and bronchoalveolar lavage fluid from mechanically ventilated intensive care unit patients

Candida albicans often resides in the oral cavity of healthy humans as a harmless commensal organism. This opportunistic fungus can cause significant disease in critically ill patients, such as those undergoing mechanical ventilation in the intensive care unit (ICU) having compromised local airway defense mechanisms. The goal of this study was to determine the intra- and inter-patient genetic relationship between strains of C. albicans recovered from dental plaque, tracheal secretions, and the lower airway by bronchoalveolar lavage of patients undergoing mechanical ventilation. Three pulsed-field gel electrophoresis (PFGE) typing methods were used to determine the genetic relatedness of the C. albicans strains, including electrophoretic karyotyping (EK) and restriction endonuclease analysis of the genome using SfiI (REAG-S) and BssHII (REAG-B). The C. albicans isolates from dental plaque and tracheo-bronchial sites from the same patient were genetically indistinguishable and retained over time, whereas strains from different patients usually separated into different genotypes. Among the three methods, REAG-B proved to be the most discriminatory method to differentiate isolates. The finding of genetically similar strains from the oral and tracheo-bronchial sites from the same patient supports the notion that the oral cavity may serve as an important source for C. albicans spread to the trachea and lung of mechanically ventilated patients

Monitoring the Long-Term Molecular Epidemiology of the Pneumococcus and Detection of Potential ‘Vaccine Escape’ Strains

While the pneumococcal protein conjugate vaccines reduce the incidence in invasive pneumococcal disease (IPD), serotype replacement remains a major concern. Thus, serotype-independent protection with vaccines targeting virulence genes, such as PspA, have been pursued. PspA is comprised of diverse clades that arose through recombination. Therefore, multi-locus sequence typing (MLST)-defined clones could conceivably include strains from multiple PspA clades. As a result, a method is needed which can both monitor the long-term epidemiology of the pneumococcus among a large number of isolates, and analyze vaccine-candidate genes, such as pspA, for mutations and recombination events that could result in 'vaccine escape' strains.We developed a resequencing array consisting of five conserved and six variable genes to characterize 72 pneumococcal strains. The phylogenetic analysis of the 11 concatenated genes was performed with the MrBayes program, the single nucleotide polymorphism (SNP) analysis with the DNA Sequence Polymorphism program (DnaSP), and the recombination event analysis with the recombination detection package (RDP).The phylogenetic analysis correlated with MLST, and identified clonal strains with unique PspA clades. The DnaSP analysis correlated with the serotype-specific diversity detected using MLST. Serotypes associated with more than one ST complex had a larger degree of sequence polymorphism than a serotype associated with one ST complex. The RDP analysis confirmed the high frequency of recombination events in the pspA gene.The phylogenetic tree correlated with MLST, and detected multiple PspA clades among clonal strains. The genetic diversity of the strains and the frequency of recombination events in the mosaic gene, pspA were accurately assessed using the DnaSP and RDP programs, respectively. These data provide proof-of-concept that resequencing arrays could play an important role within research and clinical laboratories in both monitoring the molecular epidemiology of the pneumococcus and detecting 'vaccine escape' strains among vaccine-candidate genes

Streptococcus pneumoniae Serotype-2 Childhood Meningitis in Bangladesh: A Newly Recognized Pneumococcal Infection Threat

BACKGROUND: Streptococcus pneumoniae is a leading cause of meningitis in countries where pneumococcal conjugate vaccines (PCV) targeting commonly occurring serotypes are not routinely used. However, effectiveness of PCV would be jeopardized by emergence of invasive pneumococcal diseases (IPD) caused by serotypes which are not included in PCV. Systematic hospital based surveillance in Bangladesh was established and progressively improved to determine the pathogens causing childhood sepsis and meningitis. This also provided the foundation for determining the spectrum of serotypes causing IPD. This article reports an unprecedented upsurge of serotype 2, an uncommon pneumococcal serotype, without any known intervention. METHODS AND FINDINGS: Cases with suspected IPD had blood or cerebrospinal fluid (CSF) collected from the beginning of 2001 till 2009. Pneumococcal serotypes were determined by capsular swelling of isolates or PCR of culture-negative CSF specimens. Multicenter national surveillance, expanded from 2004, identified 45,437 patients with suspected bacteremia who were blood cultured and 10,618 suspected meningitis cases who had a lumber puncture. Pneumococcus accounted for 230 culture positive cases of meningitis in children <5 years. Serotype-2 was the leading cause of pneumococcal meningitis, accounting for 20.4% (45/221; 95% CI 15%-26%) of cases. Ninety eight percent (45/46) of these serotype-2 strains were isolated from meningitis cases, yielding the highest serotype-specific odds ratio for meningitis (29.6; 95% CI 3.4-256.3). The serotype-2 strains had three closely related pulsed field gel electrophoresis types. CONCLUSIONS: S. pneumoniae serotype-2 was found to possess an unusually high potential for causing meningitis and was the leading serotype-specific cause of childhood meningitis in Bangladesh over the past decade. Persisting disease occurrence or progressive spread would represent a major potential infection threat since serotype-2 is not included in PCVs currently licensed or under development

Persistent High Burden of Invasive Pneumococcal Disease in South African HIV-Infected Adults in the Era of an Antiretroviral Treatment Program

Highly active antiretroviral treatment (HAART) programs have been associated with declines in the burden of invasive pneumococcal disease (IPD) in industrialized countries. The aim of this study was to evaluate trends in IPD hospitalizations in HIV-infected adults in Soweto, South Africa, associated with up-scaling of the HAART program from 2003 to 2008.Laboratory-confirmed IPD cases were identified from 2003 through 2008 through an existing surveillance program. The period 2003-04 was designated as the early-HAART era, 2005-06 as the intermediate-HAART era and 2007-08 as the established-HAART era. The incidence of IPD was compared between the early-HAART and established-HAART eras in HIV-infected and-uninfected individuals.A total of 2,567 IPD cases among individuals older than 18 years were reported from 2003 through 2008. Overall incidence of IPD (per 100,000) did not change during the study period in HIV-infected adults (207.4 cases in the early-HAART and 214.0 cases in the established-HAART era; p = 0.55). IPD incidence, actually increased 1.16-fold (95% CI: 1.01; 1.62) in HIV-infected females between the early-and established-HAART eras (212.1 cases and 246.2 cases, respectively; p = 0.03). The incidence of IPD remained unchanged in HIV-uninfected adults across the three time periods.Despite a stable prevalence of HIV and the increased roll-out of HAART for treatment of AIDS patients in our setting, the burden of IPD has not decreased among HIV-infected adults. The study indicates a need for ongoing monitoring of disease and HAART program effectiveness to reduce opportunistic infections in African adults with HIV/AIDS, as well as the need to consider alternate strategies including pneumococcal conjugate vaccine immunization for the prevention of IPD in HIV-infected adults

Prediction Tools for Unfavourable Outcomes in Clostridium difficile Infection: A Systematic Review

CONTEXT: Identifying patients at risk for adverse outcomes of Clostridium difficile infection (CDI), including recurrence and death, will become increasingly important as novel therapies emerge, which are more effective than traditional approaches but very expensive. Clinical prediction rules (CPRs) can improve the accuracy of medical decision-making. Several CPRs have been developed for CDI, but none has gained a widespread acceptance. METHODS: We systematically reviewed studies describing the derivation or validation of CPRs for unfavourable outcomes of CDI, in medical databases (Medline, Embase, PubMed, Web of Science and Cochrane) and abstracts of conferences. RESULTS: Of 2945 titles and abstracts screened, 13 studies on the derivation of a CPR were identified: two on recurrences, five on complications (including mortality), five on mortality alone and one on response to treatment. Two studies on the validation of different severity indices were also retrieved. Most CPRs were developed as secondary analyses using cohorts assembled for other purposes. CPRs presented several methodological limitations that could explain their limited use in clinical practice. Except for leukocytosis, albumin and age, there was much heterogeneity in the variables used, and most studies were limited by small sample sizes. Eight models used a retrospective design. Only four studies reported the incidence of the outcome of interest, even if this is essential to evaluate the potential usefulness of a model in other populations. Only five studies performed multivariate analyses to adjust for confounders. CONCLUSIONS: The lack of weighing variables, of validation, calibration and measures of reproducibility, the weak validities and performances when assessed, and the absence of sensitivity analyses, all led to suboptimal quality and debatable utility of those CPRs. Evidence-based tools developed through appropriate prospective cohorts would be more valuable for clinicians than empirically-developed CPRs

Population-Based Surveillance for Invasive Pneumococcal Disease in Homeless Adults in Toronto

BACKGROUND: Identification of high-risk populations for serious infection due to S. pneumoniae will permit appropriately targeted prevention programs. METHODS: We conducted prospective, population-based surveillance for invasive pneumococcal disease and laboratory confirmed pneumococcal pneumonia in homeless adults in Toronto, a Canadian city with a total population of 2.5 M, from January 1, 2002 to December 31, 2006. RESULTS: We identified 69 cases of invasive pneumococcal disease and 27 cases of laboratory confirmed pneumococcal pneumonia in an estimated population of 5050 homeless adults. The incidence of invasive pneumococcal disease in homeless adults was 273 infections per 100,000 persons per year, compared to 9 per 100,000 persons per year in the general adult population. Homeless persons with invasive pneumococcal disease were younger than other adults (median age 46 years vs 67 years, P<.001), and more likely than other adults to be smokers (95% vs. 31%, P<.001), to abuse alcohol (62% vs 15%, P<.001), and to use intravenous drugs (42% vs 4%, P<.001). Relative to age matched controls, they were more likely to have underlying lung disease (12/69, 17% vs 17/272, 6%, P = .006), but not more likely to be HIV infected (17/69, 25% vs 58/282, 21%, P = .73). The proportion of patients with recurrent disease was five fold higher for homeless than other adults (7/58, 12% vs. 24/943, 2.5%, P<.001). In homeless adults, 28 (32%) of pneumococcal isolates were of serotypes included in the 7-valent conjugate vaccine, 42 (48%) of serotypes included in the 13-valent conjugate vaccine, and 72 (83%) of serotypes included in the 23-valent polysaccharide vaccine. Although no outbreaks of disease were identified in shelters, there was evidence of clustering of serotypes suggestive of transmission of pathogenic strains within the homeless population. CONCLUSIONS: Homeless persons are at high risk of serious pneumococcal infection. Vaccination, physical structure changes or other program to reduce transmission in shelters, harm reduction programs to reduce rates of smoking, alcohol abuse and infection with bloodborne pathogens, and improved treatment programs for HIV infection may all be effective in reducing the risk

Non-capsulated and capsulated Haemophilus influenzae in children with acute otitis media in Venezuela: a prospective epidemiological study

<p>Abstract</p> <p>Background</p> <p>Non-typeable <it>Haemophilus influenzae </it>(NTHi) and <it>Streptococcus pneumoniae </it>are major causes of bacterial acute otitis media (AOM). Data regarding AOM are limited in Latin America. This is the first active surveillance in a private setting in Venezuela to characterize the bacterial etiology of AOM in children < 5 years of age.</p> <p>Methods</p> <p>Between December 2008 and December 2009, 91 AOM episodes (including sporadic, recurrent and treatment failures) were studied in 87 children enrolled into a medical center in Caracas, Venezuela. Middle ear fluid samples were collected either by tympanocentesis or spontaneous otorrhea swab sampling method. Standard laboratory and microbiological techniques were used to identify bacteria and test for antimicrobial resistance. The results were interpreted according to Clinical Laboratory Standards Institute (CLSI) 2009 for non-meningitis isolates. All statistical analyses were performed using SAS 9.1 and Microsoft Excel (for graphical purposes).</p> <p>Results</p> <p>Overall, bacteria were cultured from 69.2% (63 of the 91 episodes); at least one pathogen (<it>S. pneumoniae, H. influenzae, S. pyogenes </it>or <it>M. catarrhalis</it>) was cultured from 65.9% (60/91) of episodes. <it>H. influenzae </it>(55.5%; 35/63 episodes) and <it>S. pneumoniae </it>(34.9%; 22/63 episodes) were the most frequently reported bacteria. Among <it>H. influenzae </it>isolates, 62.9% (22/35 episodes) were non-capsulated (NTHi) and 31.4% (11/35 episodes) were capsulated including types d, a, c and f, across all age groups. Low antibiotic resistance for <it>H. influenzae </it>was observed to amoxicillin/ampicillin (5.7%; 2/35 samples). NTHi was isolated in four of the six <it>H. influenzae </it>positive samples (66.7%) from recurrent episodes.</p> <p>Conclusions</p> <p>We found <it>H. influenzae </it>and <it>S. pneumoniae </it>to be the main pathogens causing AOM in Venezuela. Pneumococcal conjugate vaccines with efficacy against these bacterial pathogens may have the potential to maximize protection against AOM.</p

Whole genome sequencing to investigate the emergence of clonal complex 23 Neisseria meningitidis serogroup Y disease in the United States

In the United States, serogroup Y, ST-23 clonal complex Neisseria meningitidis was responsible for an increase in meningococcal disease incidence during the 1990s. This increase was accompanied by antigenic shift of three outer membrane proteins, with a decrease in the population that predominated in the early 1990s as a different population emerged later in that decade. To understand factors that may have been responsible for the emergence of serogroup Y disease, we used whole genome pyrosequencing to investigate genetic differences between isolates from early and late N. meningitidis populations, obtained from meningococcal disease cases in Maryland in the 1990s. The genomes of isolates from the early and late populations were highly similar, with 1231 of 1776 shared genes exhibiting 100% amino acid identity and an average πN = 0.0033 and average πS = 0.0216. However, differences were found in predicted proteins that affect pilin structure and antigen profile and in predicted proteins involved in iron acquisition and uptake. The observed changes are consistent with acquisition of new alleles through horizontal gene transfer. Changes in antigen profile due to the genetic differences found in this study likely allowed the late population to emerge due to escape from population immunity. These findings may predict which antigenic factors are important in the cyclic epidemiology of meningococcal disease

A point mutation in cpsE renders Streptococcus pneumoniae nonencapsulated and enhances its growth, adherence and competence.

BACKGROUND: The polysaccharide capsule is a major virulence factor of the important human pathogen Streptococcus pneumoniae. However, S. pneumoniae strains lacking capsule do occur. RESULTS: Here, we report a nasopharyngeal isolate of Streptococcus pneumoniae composed of a mixture of two phenotypes; one encapsulated (serotype 18C) and the other nonencapsulated, determined by serotyping, electron microscopy and fluorescence isothiocyanate dextran exclusion assay.By whole genome sequencing, we demonstrated that the phenotypes differ by a single nucleotide base pair in capsular gene cpsE (C to G change at gene position 1135) predicted to result in amino acid change from arginine to glycine at position 379, located in the cytoplasmic, enzymatically active, region of this transmembrane protein. This SNP is responsible for loss of capsule production as the phenotype is transferred with the capsule operon. The nonencapsulated variant is superior in growth in vitro and is also 117-fold more adherent to and more invasive into Detroit 562 human epithelial cells than the encapsulated variant.Expression of six competence pathway genes and one competence-associated gene was 11 to 34-fold higher in the nonencapsulated variant than the encapsulated and transformation frequency was 3.7-fold greater. CONCLUSIONS: We identified a new single point mutation in capsule gene cpsE of a clinical S. pneumoniae serotype 18C isolate sufficient to cause loss of capsule expression resulting in the co-existence of the encapsulated and nonencapsulated phenotype. The mutation caused phenotypic changes in growth, adherence to epithelial cells and transformability. Mutation in capsule gene cpsE may be a way for S. pneumoniae to lose its capsule and increase its colonization potential