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PubMed2006

Rapid, sensitive, and discriminating identification of Naegleria spp. by real-time PCR and melting-curve analysis.

The free-living amoeboflagellate genus Naegleria includes one pathogenic and two potentially pathogenic species (Naegleria fowleri, Naegleria italica, and Naegleria australiensis) plus numerous benign organisms. Monitoring of bathing water, water supplies, and cooling systems for these pathogens requires a timely and reliable method for identification, but current DNA sequence-based methods identify only N. fowleri or require full sequencing to identify other species in the genus. A novel closed-tube method for distinguishing thermophilic Naegleria species is presented, using a single primer set and the DNA intercalating dye SYTO9 for real-time PCR and melting-curve analysis of the 5.8S ribosomal DNA gene and flanking noncoding spacers (ITS1, ITS2). Collection of DNA melting data at close temperature intervals produces highly informative melting curves with one or more recognizable melting peaks, readily distinguished for seven Naegleria species and the related Willaertia magna. Advantages over other methods used to identify these organisms include its comprehensiveness (encompassing all species tested to date), simplicity (no electrophoresis required to verify the product), and sensitivity (unambiguous identification from DNA equivalent to one cell). This approach should be applicable to a wide range of microorganisms of medical importance.

Animals↗
PubMed2002

Bacterial induction of autoantibodies to beta2-glycoprotein-I accounts for the infectious etiology of antiphospholipid syndrome.

The antiphospholipid syndrome (APS) is characterized by the presence of pathogenic autoantibodies against beta2-glycoprotein-I (beta2GPI). The factors causing production of anti-beta2GPI remain unidentified, but an association with infectious agents has been reported. Recently, we identified a hexapeptide (TLRVYK) that is recognized specifically by a pathogenic anti-beta2GPI mAb. In the present study we evaluated the APS-related pathogenic potential of microbial pathogens carrying sequences related to this hexapeptide. Mice immunized with a panel of microbial preparations were studied for the development of anti-beta2GPI autoantibodies. IgG specific to the TLRVYK peptide were affinity purified from the immunized mice and passively infused intravenously into naive mice at day 0 of pregnancy. APS parameters were evaluated in the infused mice on day 15 of pregnancy. Following immunization, high titers of antipeptide [TLRVYK] anti-beta2GPI Ab's were observed in mice immunized with Haemophilus influenzae, Neisseria gonorrhoeae, or tetanus toxoid. The specificity of binding to the corresponding target molecules was confirmed by competition and immunoblot assays. Naive mice infused with the affinity-purified antipeptide Ab's had significant thrombocytopenia, prolonged activated partial thromboplastin time and elevated percentage of fetal loss, similar to a control group of mice immunized with a pathogenic anti-beta2GPI mAb. Our study establishes a mechanism of molecular mimicry in experimental APS, demonstrating that bacterial peptides homologous with beta2GPI induce pathogenic anti-beta2GPI Ab's along with APS manifestations.

Animals↗
PubMed1999

Immune response to Fusobacterium nucleatum and Prevotella intermedia in patients with chronic maxillary sinusitis.

Our objective was to study the immune response to 2 anaerobic bacteria in patients with chronic maxillary sinusitis. Immunoglobulin G class antibody titers to Fusobacterium nucleatum and Prevotella intermedia were measured by enzyme-linked immunosorbent assay. Serum levels in the patients were determined on the day sinus aspiration for culture was done, and 75 to 90 days later, after completion of antimicrobial therapy, Prevotella intermedia was isolated from 16 aspirates, and F. nucleatum from 12. Median antibody levels at day 1 for F. nucleatum and P. intermedia were significantly higher in patients whose serum aspirates harbored these organisms than at days 75 to 90 in those who were cured (p < .05). However, the levels stayed elevated in the 5 patients who did not improve. The elevated antibody levels to F. nucleatum and P. intermedia, known oral pathogens, suggest a potential pathogenic role for these organisms in chronic maxillary sinusitis.

Adolescent↗
PubMed2006

Early progressive encephalopathy in boys and MECP2 mutations.

MECP2 mutations mainly occur in females with Rett syndrome. Mutations have been described in 11 boys with progressive encephalopathy: seven of nine with affected sisters and two de novo. The authors report four de novo occurrences: three pathogenic and one potentially pathogenic. Common features include failure to thrive, respiratory insufficiency, microcephaly, and abnormal motor control. MECP2 mutations should be assessed in boys with progressive encephalopathy and one or more of respiratory insufficiency, abnormal movements or tone, and intractable seizures.

Cerebral Cortex↗
PubMed1999

[Ventilator-associated pneumonia; controversies with respect to diagnosis, pathogenesis, therapy and prevention].

Ventilator-associated pneumonia (VAP) is the most frequent nosocomial infection among intensive care patients; it is associated with increased morbidity and mortality. VAP is always preceded by colonization of the upper respiratory tract with potentially pathogenic micro-organisms. Oropharyngeal colonization is pivotal in the pathogenesis of VAP, while gastric and intestinal colonization appear to be less important than generally believed. The diagnosis is difficult and usually relies on a combination of clinical, microbiological and radiographic criteria. This combination of criteria may have a high sensitivity for VAP, but specificity is low. As a result, many patients receive antibiotics unnecessarily. Bronchoscopic sampling of lower airways can increase specificity, but whether these relatively expensive techniques are cost-effective remains to be established. The best antibiotic therapy for VAP is unknown. General infection control measures remain the cornerstone of infection prevention in each intensive care unit (ICU). Selective digestive decontamination (SDD) was associated with a reduction in the incidence of VAP, but mortality rates remained largely unaffected, and selection of antibiotic-resistant pathogens remains a potential disadvantage. Routine SDD in ICU is discouraged. Decontamination of the oropharynx appears to be equally effective.

Antibiotic Prophylaxis↗
PubMed2000

The role of bacteria as a local defence mechanism in the ear, nose and throat.

The mucosae of the oro-nasopharynx in man are asymptomatically colonised by a commensal flora. This commensal flora consists largely of non-pathogenic bacteria but potentially pathogenic bacteria, such as S. pneumoniae, H. influenzae, M. catarrhalis can also be part of it. The commensal flora can be affected by host factors such as age or antibiotic usage but will in itself also affect its host. In addition to being a source of resistance genes it will also protect the host against exogenous, non-commensal pathogens. This protective effect is the result of three characteristics of the commensal flora. The commensal flora will hinder the establishment of new pathogens on the mucosa (termed colonisation resistance), it will especifically stimulate the immune system and it will induce formation of protective antibodies.

Bacteria, Aerobic↗
PubMed1997

[Prevalence of susceptibility to quinolones and other antibiotics in microorganisms isolated from community bacteriuria in Madrid in 1995. Study Group of Infections in Primary Care].

BACKGROUND: For the empiric therapy of urinary tract infections (UTI) in the extra-hospital setting the susceptibility patterns of potential pathogens, among some other factors, should be considered. Thus, the evolution in susceptibility, mainly influenced by the previous consume of antibiotic, can be observed when comparing with susceptibility patterns in the previous years. MATERIAL AND METHODS: All isolates obtained from patients with bacteriuria from 4 Health Areas in Madrid for a 3-week period were collected. The susceptibility to 4 quinolones and other 9 antibiotics was determined by the agar dilution method. The chi square test was used to compare the results obtained with Escherrichia coli isolates with those obtained with the same method in 1992. RESULTS: A total of 945 isolates were recovered. E. coli accounted for 71.4% of isolates; other gram-negative bacilli for 20.2% and gram-positive cocci for 8.4%. The percentages of E. coli isolates susceptible to ampicillin, cotrimoxazole, norfloxacin and ciprofloxacin, phosphomycin, cefuroxime, and cefuroxime-axetil were 40.5, 74.7, 91.6, 97.8, 98.1 and 75.9, respectively. Isolates with cefotaxime MIC higher than 0.5 mg/l were not detected. Significant differences were observed (p < 0.01) for susceptibilities recorded in 1992 and 1995 for amoxycillin/clavulanic acid, cefazolin, and the four quinolones. The antibiotics with the best in vitro activity against the remaining gram-negative bacilli were norfloxacin and ciprofloxacin (98% of susceptible isolates), gentamicin (97%) and cefotaxime (95%). CONCLUSIONS: Changes in susceptibility results for E. coli between 1992 and 1995 have been observed. By 1995, there are commercially available antibiotics with good in vitro activity against pathogens with the potential of causing extra-hospital UTI in our setting.

Anti-Bacterial Agents↗
PubMed2006

Enhanced identification of viral and atypical bacterial pathogens in lower respiratory tract samples with nucleic acid amplification tests.

The advantages of nucleic acid amplification tests (NAT) over conventional methods for the detection of pathogens in lower respiratory tract samples have not been established. NAT for respiratory pathogens were performed on 439 endotracheal tube (ETT) and bronchoalveolar lavage (BAL) samples. A potential pathogen was detected in 87 samples. Of 22 samples that tested positive by conventional methods, 15 tested positive for the same pathogen by NAT, 1 tested positive for a different pathogen, 2 had co-infections identified only by NAT, and 4 tested negative by NAT. An additional 73 pathogens were detected by NAT in 65 samples including 30 pathogens that were missed by conventional methods (19 adenovirus, 6 respiratory syncytial virus, 3 parainfluenza virus 1-4, 2 influenza A), 41 pathogens not routinely identified by conventional methods in most laboratories (23 rhinovirus, 8 human coronavirus OC43, 5 human metapneumovirus (hMPV), 2 human coronavirus 229E, 2 human coronavirus NL63, 1 Chlamydophila pneumoniae) and 2 pathogens from samples where no respiratory virus testing was requested (1 influenza A, 1 parainfluenza virus). Four of 52 patients who had multiple BAL samples submitted on the same day had negative and positive results by NAT on different samples. NAT improves detection of potential pathogens from ETT and BAL samples.

Adolescent↗
PubMed2001

Nasal and cloacal bacteria in free-ranging desert tortoises from the western United States.

Aerobic bacteria were collected from three free-ranging desert tortoise (Gopherus agassizii) populations in the eastern Mojave Desert (Arizona, Utah; USA) from 1989 to 1993, and from two free-ranging populations in the central Sonoran Desert (Arizona, USA) from 1990 to 1994. Six species of nasal bacteria and 18 species of cloacal bacteria were identified. At least one potential pathogen was found in the nasal cavity (Pasteurella testudinis), and at least two potential pathogens in the cloaca (Pseudomonas spp., Salmonella spp.).

Animals↗
PubMed1997

Infections in hospitalised Russian children with AIDS.

OBJECTIVES: (1) To determine the most common infections in hospitalised children with AIDS. (2) To identify the pathogens causing the above infections. (3) To investigate the possibility of nosocomial transmission of potential pathogens in a Russian AIDS hospital. SETTING: Russian AIDS Hospital for Children. SUBJECTS AND METHODS: Swabs/samples of skin, mouth, nose, urine and stool were collected from 95 hospitalised children with AIDS, 12 of their mothers and 11 health care workers in contact with the children. Hospital objects were also sampled. The predominant isolates from the infection sites were speciated. Antibiotic sensitivity profiles, phagotyping and gel electrophoresis of cell lysates were used for the epidemiological marking of Candida albicans, Staphylococcus aureus and Klebsiella pneumoniae. RESULTS: Oral infections were the most prevalent infections in this study group. The most frequently isolated micro-organisms were C. albicans, S. aureus, K. pneumoniae and Escherichia coli. The results of the epidemiological study suggests nosocomial transmission of these strains. CONCLUSIONS: The present chemoprophylaxis programme for opportunistic oral infections requires revision. Systems of microbiological surveillance and cross-infection control monitoring in relation to immunocompromised patients need to be developed to decrease the possibility of nosocomial transmission of potential pathogens between hospitalised children with AIDS, their mothers and health care providers.

AIDS-Related Opportunistic Infections↗
PubMed1984

Isolation of pathogenic bacteria from the nasopharynx of children with respiratory syncytial virus infection. Predictive value of chest roentgen examination and laboratory tests.

Potentially pathogenic bacteria were isolated from the nasopharynx in 33/66 hospitalized infants and children with verified respiratory syncytial virus (RSV) infection. The value of chest roentgenograms and blood counts for the prediction of concomitant bacterial infection was evaluated. Abnormal chest roentgenograms were found in 89% whether pathogenic bacteria were present or not. The most common finding was general hyperinflation of the lungs, with or without infiltrates. This occurred more frequently in the group with isolated RSV infection. Fever, leucocytosis and an elevated ESR were found during the illness in more than half the cases. These findings were more frequent in children who harboured potential pathogenic bacteria, though the difference was not significant. For optimal evaluation of a case all available information about it should be taken into consideration.

Bacterial Infections↗
PubMed1979

Bacteriological monitoring of unheated human milk.

To assess the bacteriological quality of unpooled expressed breast milk, a pilot bottle sample of each donation was examined before the milk was given to the neonate. Provided the milk did not contain greater than 2500 organisms/ml or potential pathogens it was used unheated. Milk containing between 2500 and 5000 organisms/ml and no potential pathogens was used after pasteurisation. Using these criteria, 67% of 460 donations were acceptable. However, because the bacteriological quality varied, 45% of domiciliary donations were discarded compared with only 29% of those from hospital.

Bacteriological Techniques↗
PubMed1977

An extensive rotavirus outbreak in aboriginal infants in central Australia.

Faecal specimens were obtained from 92 Aboriginal infants admitted to Alice Springs Hospital during an extensive outbreak of gastroenteritis in the surrounding area in July, 1976. Specimens were examined for potentially pathogenic viruses, bacteria and parasites. Rotavirus proved to be by far the most common potential pathogen, being detected in 54% of specimens.

Australia↗
PubMed1999

Development of digoxigenin-labeled PCR amplicon probes for use in the detection and identification of enteropathogenic Yersinia and Shiga toxin-producing Escherichia coli from foods.

By including digoxigenin-11-dUTP in a polymerase chain reaction (PCR), amplification products were produced that contained nonisotopic markers for use as DNA hybridization probes. Because these labeled amplicons encode pathogenic traits for specific foodborne bacteria, they can be used to detect the presence of potentially virulent organisms that may be present in foods. This technology allows the synthesis of a variety of shelf-stable probe reagents for detecting a number of foodborne microbes of public health concern. We used this technology to detect four genes in two potential pathogens: virF and yadA in enteropathogenic Yersinia and stx1 and stx2 in Shiga-like toxin-producing Escherichia coli. Results of DNA hybridizations of dot blots of 68 Yersinia strains and 24 of 25 E. coli strains were consistent with results of equivalent PCR analyses. DNA colony hybridization with nonisotopic virF probes of colonies arising on spread plates from artificially contaminated food homogenates was able to detect potentially pathogenic Y. enterocolitica. When compared with oligonucleotide probes, amplicon probes are much less sensitive to changes in hybridization and wash temperatures, allowing greater reproducibility. Labeled probe preparations were reused more than five times and have been stored at -20 degrees C for more than 8 months. This method conveniently generates probes that are safe, stable, inexpensive, reusable, and reliable.

Adhesins, Bacterial↗
PubMed2004

Intra-host evolution of human immunodeficiency virus type 1 and viral fitness.

RNA viruses are frequently tolerant to high levels of mutagenesis. In contrast, DNA viruses are less errorprone and coevolve along with their specific hosts over long time periods. Although both strategies have been successful, the "RNA-strategy" (directly linked to the pathogenic potential of these agents) most often generates novelty (new variants, new strains, and even new viral pathogens). For several decades, intra-host virus evolution has been considered to be a speculative field, far from the main issues of clinical virology. This concept is now changed, due to the evidence that RNA virus evolution is intimately linked to failures in viral disease control and prevention. Antiviral strategies using single and fixed elements (i.e. treatments using one antiviral compound, immunizations using a single recombinant protein) have been unable to control highly dynamic quasispecies, such as human immunodeficiency virus type I (HIV-1) and hepatitis C virus (HCV). The development of combinatorial treatments in HIV-1 infection and the recognition that vaccines should be multivalent are important steps in adapting disease control strategies to the complexity of viral populations. The present report summarizes the strategies adopted to address HIV-1 evolution and its phenotypic consequences, including changes in susceptibility to antiviral compounds, viral fitness, and pathogenic potential. In particular, it is highlighted that sequence-function analyses of the intra-host HIV-I evolution, including studies of viral fitness, have opened up new perspectives not only to studying the pathogenic mechanisms and the virus-host relationships, but also to designing new strategies for monitoring antiviral therapies.

AIDS Vaccines↗
PubMed2003

Prepared endotracheal tubes: are they a potential source for pathogenic microorganisms?

UNLABELLED: Prepared endotracheal tubes (PETTs) are frequently used for unanticipated difficult intubation, but their storage time is highly variable and institution-dependent. We sought to determine first, if open, unused PETTs are a potential source of pathogenic microorganisms, and second, if PETTs can provide a medium for bacterial survival after deliberate contamination. A stylet was inserted into a 7-mm ETT, and this system was ethylene oxide sterilized. The PETTs were placed in 20 different locations and sampled 8 times in a 4-wk period. Growth was determined after 48-h incubation, and the microorganism was identified. In Phase 2, the PETT (n = 40) was swabbed with a fresh suspension of H. influenzae, Pseudomonas aeruginosa, Staphylococcus aureus, Enterococcus faecium, or a negative control. Nonvirulent bacteria were cultured from 13 of 160 (8.1%) samples and from 15 of 320 (4.7%) samples in Phases 1 and 2, respectively. No PETT grew the same bacteria more than once. In Phase 2, after 24 h, only E. faecium was recovered. Based on this study, the pathogenic potential of PETTs is very small, and they can be safely used for up to 1 mo. This practice could translate to significant cost reduction for operating room budgets. IMPLICATIONS: Prepared endotracheal tubes (PETTs) are back-up airway equipment to be used in the case of a difficult intubation. A short PETT shelf life because of unknown safe storage time results in significant budget costs. This blinded, controlled study examined the pathogenic potential of PETTs in the operating room environment.

Bacteria↗
PubMed2003

Microbial dynamics of purulent nasopharyngitis in children.

This review presents the microbiological dynamic and therapeutic options in the management of purulent nasopharyngitis (NPT). The nasopharynx (NP) of healthy children is generally colonized by relatively non-pathogenic aerobic and anaerobic organisms, some of, which possess the ability to interfere with the growth of potential pathogens. Conversely, carriage of potential respiratory aerobic pathogen such as Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis, as well as some anaerobic bacteria (Peptostreptococcus, Fusobacterium and Prevotella spp.) increases during purulent NPT. The development of purulent NPT in children is associated with the pre-existing colonization by potential pathogens and the absence of interfering organisms in the NP. Controversy exists regarding the management of NPT as no conclusive evidence exists to date that the administration of antimicrobials will shorten the illness.

Anti-Bacterial Agents↗
PubMed1989

Mixed infections in childhood diarrhoea: results of a community study in Kiambu District, Kenya.

One thousand four hundred and twenty diarrhoea specimens from 846 children aged 0 to 60 months were collected and analysed for bacteria, parasites and rotavirus over a 16 month period, from June 1985 to September 1986 inclusive. The study was conducted in 4 villages situated in Kiambu District, Kenya. All the specimens were analysed for rotavirus and parasites, including Cryptosporidium. The majority of the specimens were analysed for enteropathogenic Escherichia coli (EPEC), Shigella, Salmonella, Campylobacter and Aeromonas. Only 387 specimens were analysed for enterotoxigenic E. coli (ETEC). However, of this proportion analysed for ETEC, 33% were positive. A total of 344 specimens were negative for any organisms while a further 140 were only positive for parasites which have been implicated as being pathogenic, including Ascaris lumbricoides, Hymenolepis nana, Trichomonas hominis and Blastocysts hominis were considered to be at least potentially pathogenic and capable of causing diarrhoea. An average of only 29.4% of these organisms occurred as single isolates. The remaining infections were mixed, with a maximum of 7 potentially pathogenic organisms occurring together in a single specimen. The associations of certain organisms were significant, notably Campylobacter with Giardia lamblia. Campylobacter with EPEC, EPEC with Ascaris, and G. lamblia with rotavirus. The latter was a negative association.

Community Health Services↗