Diagnostic Microbiology & Infectious Disease - Articles in Press

Susceptibility to antifungal agents and genotypes of Brazilian clinical and environmental Cryptococcus gattii strains - Corrected Proof

2012-02-16

Abstract: There are few reports concerning the in vitro antifungal susceptibility of clinical and environmental Cryptococcus gattii isolates. In this study, we performed polymerase chain reaction–restriction fragment length polymorphism to investigate the molecular subtypes of 50 clinical and 4 environmental Brazilian isolates of C. gattii and assessed their antifungal susceptibility for fluconazole (FLU) and amphotericin B (Amb) according to recent recommendations proposed for antifungal susceptibility testing of nonfermentative yeasts. Time–kill curve studies were performed using RPMI 1640 medium to analyze the fungicidal effect of AmB. We found 47 VGII (94%) molecular types and 3 VGI (6%) types among the clinical isolates. The environmental isolates were VGII (75%) subtype and VGI (25%) subtype. The FLU-MIC ranged from 1 to 64 mg L−1, and MIC50/MIC90 values were, respectively, 8/16 mg L−1. For AmB, the MICs were low and homogeneous, ranging from 0.12 to 0.5 mg L−1, for VGI or VGII. The time required to reach the fungicidal end point (99.9% killing) was 6 h for the majority of strains (64%), but viable cells of VGII were still present after 48 h of exposition. We pointed out the occurrence of high FLU-MICs for C. gattii isolates with highest values for VGII. Our data also suggest that the rate of killing of C. gattii by AmB is strain dependent, and viable cells of VGII genotype strains were still observed after an extended incubation time, addressing future studies to determine whether the in vitro fungicidal activity could be clinically relevant.

Prevalence of Bordetella pertussis and Bordetella parapertussis infections in Tunisian hospitalized infants: results of a 4-year prospective study - Corrected Proof

2012-02-10

Abstract: The prevalence of pertussis in Tunisia remains undetermined essentially because of the unavailability of a basic laboratory diagnostic service. Specific diagnostic tools were applied for the first time in a Tunisian prospective study in order to get a first estimation of the prevalence of Bordetella pertussis/parapertussis infections and to evaluate their use to determine the epidemiologic characteristics of these infections in Tunisian infants. Between 2007 and 2011, a total of 626 samples from 599 infants aged <1 year with and without pertussoid cough were investigated for the presence of B. pertussis/parapertussis using culture and real-time polymerase chain reaction (PCR). The real-time PCR (RT-PCR) targets include IS481 commonly found in B. pertussis, B. bronchiseptica, and B. holmesii; IS1001 specific of B. parapertussis, in combination with the pertussis toxin promoter region gene (ptx) of B. pertussis; and the recA gene specific of B. holmesii. When possible, patients' household contacts provided nasopharyngeal aspirates (NPAs) for RT-PCR detection of B. pertussis/parapertussis or single-serum samples for anti-PT IgG quantification. All except 1 NPAs were negative by conventional culture, whereas PCR gave positive signals for 126 specimens (21%): B. pertussis, B. parapertussis, and Bordetella spp. were detected in 82%, 6%, and 4% of the samples, respectively. The simultaneous presence of B. pertussis and B. parapertussis was noted in 8% of the cases. Pertussis was reported throughout the year with a peak during the summer of the year 2009. The prevalence of Bordetella infection was 20% between 2007 and 2011. Most of these cases corresponded to patients younger than 6 months who received <3 doses of pertussis vaccine. Among the household contacts enrolled in the study, mothers seemed to be the likely source of infection. This study showed that pertussis is still prevalent in Tunisia and that the disease remains a public health problem affecting not only infants but also adults. Given this situation, sensitive and specific laboratory tests are needed to improve the accuracy of pertussis diagnosis.

Molecular characterization of Neisseria meningitidis B:NT:P1.14/162 clonal complex responsible of invasive meningococcal disease in the north of Italy - Corrected Proof

2012-02-10

Abstract: The molecular characteristics of 14 B:NT:P1.14 Neisseria meningitidis isolates, collected from 2007 through 2010 in Italy, have been investigated. The B:NT:P1.14 phenotype has only more recently been identified in our country, mainly associated with clonal complex CC162, which is rare in Italy.

Impact of yeast–bacteria coinfection on the detection of Candida sp. in an automated blood culture system - Corrected Proof

2012-02-06

Abstract: Invasive candidiasis remains a major cause of morbidity and mortality. It is now well known that an early diagnosis contributes to the patients' outcome. Blood cultures, which are the first-line test in case of bloodstream infection suspicion, can be carried out using fungus-selective medium (containing antibiotics) or standard microorganism medium allowing both bacterial and fungal growth. Some patients can suffer from polymicrobial sepsis involving bacteria and yeasts, so we decided to investigate in blood cultures the influence of the presence of bacteria on fungal development. Simulated blood cultures were performed using Candida albicans or C. glabrata coincubated with Escherichia coli or Staphylococcus aureus at different concentrations. The results showed that, in a standard microorganism medium, bacterial growth could hide the fungal development. Thus, in patients at risk of invasive candidiasis, the use of a specific fungal medium could improve the diagnosis and allow an earlier efficient antifungal treatment.

Analysis of Streptococcus pneumoniae secreted antigens by immuno-proteomic approach - Corrected Proof

2012-02-06

Abstract: Streptococcus pneumoniae is an important human pathogen that causes a variety of diseases in both adults and children, such as pneumonia, bacteremia, meningitis, otitis media, and sinusitis. Despite their clinical importance, to date, there have been few proteomic studies of these strains for screening of virulence factors or diagnostic markers. In the present study, secreted proteins (secretome) of Streptococcus pneumoniae strains were enriched using ammonium sulfate precipitation and identified by the shotgun proteomic method using 1-dimensional electrophoresis liquid chromatography–mass spectrometry/mass spectrometry analysis. Characterization of the identified proteins revealed that 17.8% (42) of the secreted proteins possessed signal peptides. Twenty-one secreted proteins belonged to the extracellular group, and 4 secreted proteins belonged to the cell wall group. Well-known virulence factors (PrtA, PspC, PsaA, PbpA, PhtD, AmiA, ZmpB, Eno, and Ply) were included in the secreted protein fraction. Western blotting using antiserum against secreted protein mixtures showed that Gsp-781, Sphtra, NagA, PhtD, ZmpB, and Eno were strongly immunogenic. Our data suggest that the immuno-proteomic approach is a useful method for high-throughput identification of secreted proteins and screening of candidate vaccine antigens or diagnostic markers. Gsp-781 is introduced as a novel secreted antigen of Streptococcus pneumoniae.

Fecal colonization of VIM-1–producing Klebsiella pneumoniae and in vivo transfer of multidrug-resistant IncN plasmid in a renal transplant patient - Corrected Proof

Umaer Naseer, Bjørn Odvar Eriksen, Arnfinn Sundsfjord, Ørjan Samuelsen — 2012-02-02

Abstract: We report a case of long-term colonization of a carbapenemase (VIM)-producing Klebsiella pneumoniae clone in a renal transplant patient and demonstrate the in vivo transmission of a broad-host-range multidrug-resistant IncN plasmid containing blaVIM, blaSHV-12, and qnrS to Escherichia coli.

The presence of genes encoding for different virulence factors in clonally related Escherichia coli that produce CTX-Ms - Corrected Proof

Akke K. Van der Bij, Gisele Peirano, André Pitondo-Silva, Johann D.D. Pitout — 2012-02-02

Abstract: Successful international clones have recently emerged among Escherichia coli that produce CTX-M β-lactamases as important causes of community-onset urinary tract and bloodstream infections. One hundred and seven isolates that belong to sequence types (STs) ST38, ST131, ST405, ST648, and 38 nonrelated CTX-M–producing E. coli from Canada and the Netherlands were assigned to phylogenetic groups and tested for the presence of genes encoding for virulence factors (VFs) using established multiplex polymerase chain reaction. The STs E. coli were significantly more resistant to antibiotics—ST38, ST405, and ST648 belonged to phylogenetic group D while ST131 belonged to B2. Secreted autotransporter toxin (sat), aerobactin receptor, and pathogenicity island marker were significantly more common among the STs; the heat-resistant agglutinin (hra) was present in ST38, sat, and uropathogenic-specific protein, and putative adhesin-siderophore receptor was more common in ST131, while outer membrane protease T was present in ST648. ST131 had a significantly higher VF score. In conclusion, the precise role of these VFs remains to be elucidated; however, we have identified certain putative VFs that possibly contribute to the fitness and success of certain sequence types.

Evaluation of the PrimerDesign™ genesig real-time reverse transcription–polymerase chain reaction assay and the INFINITI® Respiratory Viral Panel Plus assay for the detection of human metapneumovirus in Kuwait - Corrected Proof

Mariam Al-Turab, Wassim Chehadeh, Fahd Al-Mulla, Widad Al-Nakib — 2012-02-02

Abstract: Human metapneumovirus (hMPV) is a respiratory pathogen that was discovered in 2001 and is considered a major cause of both upper and lower respiratory tract infections. A sensitive, fast, and high-throughput diagnostic test is needed for the detection of hMPV that may assist in the clinical management as well as in the reduction of inappropriate therapy. Therefore, a comparison assessment was performed in this study between the PrimerDesign™ genesig real-time reverse transcription–polymerase chain reaction (RT-PCR) Assay and the INFINITI® Respiratory Viral Panel Plus Assay (RVP-Plus) for the detection of hMPV infection in patients with respiratory tract infections. A total of 200 respiratory samples were collected from 185 hospitalized patients, during the winter season in Kuwait. Of 185 patients, 10 (5.4%) were positive for hMPV RNA by the in-house RT-PCR assay, while 7 (4%) were positive for hMPV RNA by the real-time RT-PCR assay and 9 (5%) were positive for hMPV RNA by the INFINITI® RVP-Plus assay. The high incidence rate (60%) of hMPV infection was in January 2011. The sensitivity of the real-time RT-PCR and INFINITI® RVP-Plus assays was 70% and 90%, respectively, with specificity of 100% for both assays. hMPV types A and B could be identified in this study; however, discordant genotyping results were found between the direct sequencing method and the INFINITI® RVP-Plus assay in 33% of hMPV-positive patients.

Direct identification of mycobacteria from smear-positive sputum samples using an improved multiplex polymerase chain reaction assay - Corrected Proof

Ju-Hsin Chia, Tsu-Lan Wu, Lin-Hui Su, An-Jing Kuo, Hsin-Chih Lai — 2012-01-27

Abstract: The rapid identification of mycobacteria from smear-positive sputum samples is very important. To identify the Mycobacterium tuberculosis complex (MTBC) and frequently isolated nontuberculous mycobacteria strains directly from smear-positive sputum samples, an improved multiplex polymerase chain reaction (PCR) assay was developed. Nine pairs of primers targeting the 16S–23S rDNA internal transcribed spacer-1, hsp65, and the early secretory antigen (ESAT-6) gene sequences were developed, and their efficacy was evaluated in comparison to traditional culturing and 16S rRNA gene sequencing methods. A total of 200 smear- and culture-positive sputum specimens collected between November 2005 and May 2006 were used for the analysis. The results of the assay showed an accurate identification rate for acid-fast bacilli (AFB) 3+, AFB 2+, and AFB rare/1+ samples of 98%, 95%, and 53%, respectively. The improved multiplex PCR method saves time and has advantages for identifying mycobacteria from AFB 2+ and 3+ sputum samples. The method is suitable for use in countries with a high MTBC prevalence rate.

Galactomannan testing might be useful for early diagnosis of fusariosis - Corrected Proof

2012-01-27

Abstract: Galactomannan (GM) is used to diagnose aspergillosis. We present a case of a hematopoietic stem cell transplantation recipient with fusariosis who received early antifungal treatment based on GM positivity. Additionally, 3 Fusarium isolates tested positive for GM. Fusarium is another mold containing GM. GM might be useful for diagnosing fusariosis in high-risk patients.

Efficiency of the TK Culture System in the diagnosis of tuberculosis - Corrected Proof

2012-01-25

Abstract: We have evaluated the efficiency of the TK Rapid Mycobacterial Culture System in isolating mycobacteria from clinical samples and in susceptibility testing. The TK Medium indicates mycobacterial growth by changing its color from red to yellow. During a 1-year period, 16,303 clinical samples were inoculated to TK selective (TK SLC) and Löwenstein–Jensen media (LJ). Mycobacteria were isolated in 2150 (13.04%) samples in at least 1 type of medium. While LJ isolated mycobacteria from 1920 (11.69%) of all samples, TK SLC isolated 2070 (12.63%). Among all positives, the isolation rates for LJ and TK SLC were 89.30% and 96.27%, respectively. Contamination of cultures by other organisms was observed in 878 (5.33%) LJ tubes and in 90 (0.55%) TK SLC tubes. On average, time-to-growth detection was 15.57 days in TK SLC and 25.14 days in LJ. The modes of time-to-growth detection were 12 and 25 days for TK SLC and LJ, respectively. The reliability of antimycobacterial susceptibility testing was checked by 36 Mycobacterium tuberculosis strains with known susceptibility patterns which were obtained from the World Health Organization collection and by participating in an external quality control program. All susceptibility results, except for a few borderline-resistant strains, were consistent with the expected susceptibility patterns. The TK Rapid Mycobacterial Culture System is a practical and reliable automated system that shortens the time required for both culture and susceptibility results. All types of TK Media are ready to use, saving time and effort as well as drastically reducing contamination during testing.

Apropos “Evaluation of third-generation RIDASCREEN enzyme immunoassay for the detection of norovirus antigens in stool samples of hospitalized children in Belém, Pará, Brazil” - Corrected Proof

2012-01-25

We read with interest the article of , “Evaluation of third-generation RIDASCREEN® enzyme immunoassay in stool samples of hospitalized children in Belém, Pará, Brazil.” This study would be valuable to formulate future plans to screen large numbers of clinical samples in order to address norovirus (NoV) epidemiology. Our intention is to share previously published () findings on third-generation RIDASCREEN® kit evaluation in Brazil and contribute to the knowledge in diagnosis of NoV infection.

Characterization of ST80 Panton-Valentine leukocidin-positive community-acquired methicillin-resistant Staphylococcus aureus clone in Tunisia - Corrected Proof

Mouna Ben Nejma, Maha Mastouri, Besma Bel Hadj Jrad, Mohamed Nour — 2008-04-04

Abstract: The spread of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) has been reported in communities worldwide. In this study, we characterized 64 Tunisian CA-MRSA by agr typing, polymerase chain reaction assay for 20 virulence genes, staphylococcal chromosomal cassette mec (SCCmec) typing, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and protein A gene (spa) typing. All our isolates were lukS-PV-lukF-PV positive, etd positive, and edin positive. They harbored SCCmec type IV and belonged to agr group 3. PFGE typing showed that our isolates were distributed in 11 different pulsotypes. spa typing and MLST, performed with isolates representative of each PFGE pattern, revealed that all isolates had a unique spa type (t044) and a common sequence type (ST80). The isolates showed susceptibility to the majority of antibiotics, and resistance to kanamycin, erythromycin, and tetracycline, but intermediate resistance to fusidic acid. Full analysis of our results revealed that our isolates were nonmultiresistant and belonged to a single clonal type ST80.

WITHDRAWN: Selection of a surrogate β-lactam testing agent for initial susceptibility testing of doripenem, a new carbapenem - Corrected Proof

Ronald N. Jones, Helio S. Sader, Thomas R. Fritsche, Michael J. Janechek — 2007-10-05

This article has been withdrawn consistent with Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). The Publisher apologizes for any inconvenience this may cause.

WITHDRAWN: Yersinia enterocolitica identification in stool samples using real-time PCR - Corrected Proof

Zheng HaoXuan, Wang JiDe, Zhang MingJun, Sun Yong, Jiang Bo — 2007-06-08

This article has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy