Evaluation of two rapid methods for identifying and biotyping Haemophilus influenzae.

Murphy, Philip G.; Craig, I.K.; Lafong, A C; Smyth, E.T.M.

doi:10.1136/jcp.43.7.581

Cited by 5 publications

(1 citation statement)

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“…Biochemical and phenotypic analyses of H. influenzae and H. haemolyticus traits were performed as previously described by Kilian (25,26). Briefly, the emission of H 2 S was assessed with lead acetate strips (Fisher Scien- (26,25,39,60). Briefly, indole production was assessed by placing a small amount of bacteria on filter paper saturated with Kovács reagent (Remel) and observing the paper for a purple color reaction; urease activity was assessed in rapid urea broth (BD), according to the manufacturer's instructions, by mixing an inoculum loop of bacteria from the plate growth and observing the plate for a color reaction after 4 h of incubation at 37°C; and ODC activity was determined with ODC broth (Remel), according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

Relationships of Nontypeable Haemophilus influenzae Strains to Hemolytic and Nonhemolytic Haemophilus haemolyticus Strains

et al. 2008

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Haemophilus influenzae is both a human respiratory pathogen and pharyngeal commensal, while H. haemolyticus, the closest phylogenetic relative of H. influenzae, is arguably a strict pharyngeal commensal. A hemolytic phenotype has historically differentiated H. haemolyticus from H. influenzae, but the recent recognition of significant nonhemolytic H. haemolyticus colonization has decreased this trait's resolvability. Given this and the potential of recombination between the species, we examined the distribution of microbiologic and molecular traits between collections of H. influenzae and H. haemolyticus strains separated within a dendrogram obtained by multilocus sequence analysis (MLSA). All strains hybridizing with a probe to iga, a gene encoding an immunoglobulin A protease of H. influenzae, clustered apart from strains that did not hybridize with the probe. Other traits also segregated significantly along this division, suggesting a separation of the species. Of note, the LOS genes licA, lic2A, and lgtC of H. influenzae were approximately 2, 6, and 54 times, respectively, more prevalent in H. influenzae than in H. haemolyticus. In contrast to species separation, interspecies recombination was evidenced by the inability of single gene sequences to phylogenetically separate the species and by the "fuzzy" distribution of some species-specific traits across the species dividing line. Together, these data support the historically accurate and pragmatic division of these species while recognizing their potential for recombination. Future comparative genomic studies identifying common and distinctive genes could be useful in evaluating their role in the commensal or virulent growth, respectively, of H. influenzae.Haemophilus influenzae is one of eight Haemophilus species that reside as commensal organisms in the pharyngeal cavity of humans. These other commensal Haemophilus species include H. parainfluenzae, H. haemolyticus, H. parahaemolyticus, H. paraphrohaemolyticus, H. segnis, H. aphrophilus, and H. paraphrophilus. H. influenzae is by far the most pathogenic member of the genus in humans. Strains possessing a type b capsule are often associated with invasive diseases, such as meningitis, sepsis, and pneumonia; and strains lacking a capsule (referred to as nontypeable [NT]) are associated with localized mucosal diseases, such as otitis media, sinusitis, and bronchitis. Other haemophili are considered to be rarely associated with disease, and H. haemolyticus has never been implicated as an infectious disease agent (1).In contrast to other haemophili, H. haemolyticus and H. influenzae both depend on X (hemin) and V (NAD) growth factors, both lack the ability to ferment sucrose, both share similar GϩC contents, and both are nearly indistinguishable by their colony and cellular morphologies (25). Initial phylogenetic studies performed by using DNA-DNA hybridization revealed that H. haemolyticus associated with H. influenzae in a cluster of the family Pasteurellaceae termed Haemophilus sensu stricto (6,41,47), and t...

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Section: Methodsmentioning

confidence: 99%

Relationships of Nontypeable Haemophilus influenzae Strains to Hemolytic and Nonhemolytic Haemophilus haemolyticus Strains

et al. 2008

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Haemophilus

Slack

2010

Topley &Amp; Wilson's Microbiology and Microbial Infections

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Multicenter Evaluation of the New Vitek 2 Neisseria - Haemophilus Identification Card

et al. 2008

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The new Neisseria-Haemophilus identification (NH) card for Vitek 2 was compared with 16S rRNA gene sequencing (16S) as the reference method for accurate identification of Neisseria spp., Haemophilus spp., and other fastidious gram-negative bacteria. Testing was performed on the Vitek 2 XL system with modified software at three clinical trial laboratories. Reproducibility was determined with nine ATCC quality control strains tested 20 times over a minimum of 10 days at all three sites. A challenge set of 30 strains with known identifications and 371 recent fresh and frozen clinical isolates were also tested. Expected positive and negative biochemical reactions were also evaluated for substrate reproducibility. All microorganisms were tested on the NH card, and all clinical and stock isolates were saved for 16S testing. All reproducibility tests yielded expected results within a 95% confidence interval. For challenge microorganisms, there was 98% overall correct identification, including 8% low discrimination, 2% incorrect identification, and 0% unidentified. For clinical strains, there was 96.5% overall correct identification, including 10.2% low discrimination, 2.7% incorrect identification, and 0.8% unidentified. The 2.7% (10/371) of clinical isolates that gave an incorrect identification consisted of 7 isolates correct to genus and 3 strains incorrect to genus. There were an additional 27 strains (primarily Neisseria species) for which the 16S identification result was different from the NH card result. These were all unclaimed species by the system. The new NH card met all performance criteria within a 95% confidence interval compared to identification of clinical isolates by 16S.In the diagnostic clinical microbiology laboratory, rapid and accurate identification of bacterial pathogens is essential for prompt and appropriate management of infected patients. This is important for bacterial pathogens within the genera Haemophilus and Neisseria.The science has evolved over many years. Classical methods gave way to substrate-based identification of these microorganisms involving overnight and then shorter incubation. More recently molecularly based identification has started to change the landscape (2,5,7,8,12), but for most laboratories that lack such sophisticated technical capabilities, the ability to use either a manual or automated system that will give a high level of correct identifications is sufficient for most purposes.For identification of Haemophilus and Neisseria species, a variety of commercial methods have been available on the market for many years. For Neisseria species these have included Neisseria-Kwik, Gonogen, Gonochek II, RIM-N, API QuadFERM-Plus, Minitek, Identicult-Neisseria, and several others (1,6,16,18). All of these are based on colorimetric changes in miniaturized substrates, with either enzymatic or growth end points. The ability to identify and separate Haemophilus species from nonpathogenic Neisseria species, particularly those from the respiratory tract, then led to the addition ...

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Evaluation of two rapid methods for identifying and biotyping Haemophilus influenzae.

Cited by 5 publications

References 8 publications

Relationships of Nontypeable Haemophilus influenzae Strains to Hemolytic and Nonhemolytic Haemophilus haemolyticus Strains

Relationships of Nontypeable Haemophilus influenzae Strains to Hemolytic and Nonhemolytic Haemophilus haemolyticus Strains

Haemophilus

Multicenter Evaluation of the New Vitek 2 Neisseria - Haemophilus Identification Card

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