Biomedical subjects

P H Hendolin

Publications and source records attributed to P H Hendolin.

5 recordsLinked to original sources

PubMed2002

Detection of Alloiococcus otitidis in the nasopharynx and in the outer ear canal.

Alloiococcus otitidis has been recovered from the middle ear of children with otitis media with effusion, but its natural habitat is not known. To determine whether the nasopharynx and the outer ear canals are the natural habitats of A. otitidis, 145 swabs (50, nasopharynx; 95 outer ear canal) collected from 50 children were screened by polymerase chain reaction. A. otitidis DNA was detected in seven (4.8%) of the 145 specimens, of which four were nasopharynx, and three outer ear canal. These results indicate that the nasopharynx and outer ear canal may be the body sites for localization of A. otitidis.

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PubMed2000

Clinically applicable multiplex PCR for four middle ear pathogens.

The multiplex PCR method for the detection of Alloiococcus otitidis, Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae (P. H. Hendolin, A. Markkanen, J. Ylikoski, and J. J. Wahlfors, J. Clin. Microbiol. 35:2854-2858, 1997) in middle ear effusions (MEEs) was modified to be better suited for clinical use. To detect false-negative results, an internal amplification was added to the reaction, and to prevent carryover contamination, the dUTP-uracil-N-glycosidase system was incorporated into the procedure. Labor was minimized by using the heat-activatable AmpliTaq Gold polymerase in order to circumvent manual hot start and by detecting the amplification products on an automated sequencer. The performance of the improved protocol was verified with MEEs from patients with otitis media with effusion. In addition, a ligase detection reaction (LDR) was developed for confirmation of the PCR products. The modifications increased the reliability of the protocol and the hands-off time significantly. However, when two DNA extraction protocols were compared, gram-negative bacteria were detected more often in phenol-treated MEEs (94 versus 46%; P < 0.001), and gram-positive bacteria were detected more often in MEEs dissolved in sodium dodecyl sulfate-NaOH-chaotropic salt (83 versus 27%; P < 0.001). The LDR was found to be 100% specific. In all, the results demonstrate the feasibility of the rapid (7-h) multiplex PCR method for routine laboratory use.

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PubMed2000

Panfungal PCR and multiplex liquid hybridization for detection of fungi in tissue specimens.

A procedure based on panfungal PCR and multiplex liquid hybridization was developed for the detection of fungi in tissue specimens. The PCR amplified the fungal internal transcribed spacer (ITS) region (ITS1-5.8S rRNA-ITS2). After capture with specific probes, eight common fungal pathogens (Aspergillus flavus, Aspergillus fumigatus, Candida albicans, Candida krusei, Candida glabrata, Candida parapsilosis, Candida tropicalis, and Cryptococcus neoformans) were identified according to the size of the amplification product on an automated sequencer. The nonhybridized products were identified by sequencing. The performance of the procedure was examined with 12 deep-tissue specimens and 8 polypous tissue biopsies from the paranasal sinuses. A detection level of 0.1 to 1 pg of purified DNA (2 to 20 CFU) was achieved. Of the 20 specimens, PCR was positive for 19 (95%), of which 10 (53%) were hybridization positive. In comparison, 12 (60%) of the specimens were positive by direct microscopy, but only 7 (35%) of the specimens showed fungal growth. Sequencing of the nonhybridized amplification products identified an infecting agent in six specimens, and three specimens yielded only sequences of unknown fungal origin. The procedure provides a rapid (within 2 days) detection of common fungal pathogens in tissue specimens, and it is highly versatile for the identification of other fungal pathogens.

Adult↗

PubMed1999

High incidence of Alloiococcus otitis in otitis media with effusion.

BACKGROUND: The etiology of otitis media with effusion (OME) is unclear. Although the majority of effusions show inflammation, culture methods yield positive results for bacteria in only 20 to 30% of cases. METHODS: The polymerase chain reaction was used for detection of three upper respiratory tract pathogens, Haemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumoniae, and a fairly recently described bacterium, Alloiococcus otitis (A. otitidis), that is solely found in OME. The study included 67 middle ear effusions that were collected from 48 pediatric OME patients during ventilation tube placement. RESULTS: PCR tested positive for 57 (85.1%) of the middle ear effusions. Thirty-one (46.3%) A. otitis-, 12 (17.9%) H. influenzae-, 25 (37.3%) M. catarrhalis- and 14 (20.9%) S. pneumoniae-positive effusions were obtained. All four study organisms showed similar distribution in effusions of various duration (P = 0.72) and in different effusion types (P = 0.59). Only the proportion of M. catarrhalis-positive effusions was lowered by recent antimicrobial therapy (P < 0.05). Although the study organisms had equal distributions among singly and multiply positive specimens (P = 0.90), A. otitis was detected significantly more often with one of the three other species (15 of 19, 78.9%) than the other species with each other (4 of 19, 21.1%, P < 0.001). CONCLUSIONS: The findings suggest a bacterial etiology for OME. Association of A. otitis with the three other species implies that this organism might have the capability of augmenting bacterial colonization in the middle ear.

Child↗

PubMed1997

Use of multiplex PCR for simultaneous detection of four bacterial species in middle ear effusions.

A multiplex PCR procedure was developed for the simultaneous detection of Alloiococcus otitidis, Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae in middle ear effusions (MEEs) from patients with chronic otitis media with effusion. The bacterial 16S rRNA gene was chosen as the target, and the procedure used one common lower primer and four species-specific upper primers. The reaction was optimized by changing the primer concentrations to yield equal amounts of amplification products. The specificity of the reaction was verified with various bacterial species found in the nasopharynx. The performance of the procedure was examined with 25 MEE specimens, and the results were compared to those obtained by conventional culture methods. A detection level of 10 bacterial cells/reaction for each of the study organisms was achieved. By conventional culture methods, 8 (32%) of the specimens showed growth of one of the study organisms. In contrast, 21 (84%) of the specimens tested positive by the multiplex PCR. None of the culture-positive specimens were PCR negative, whereas three (12%) of the PCR-positive specimens tested positive for two of the four study organisms. Thus, the multiplex PCR method improves the detection rate significantly compared to that of the conventional culture method.

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