Rapid Strep System Research Articles

The association of Lactococcus petauri with lactococcosis is older than expected

Lactococcosis is a globally prevalent infectious disease that has a significant economic and sanitary impact on the rainbow trout industry. Lactococcus garvieae has traditionally been considered the only species implicated in the etiology of this disease, but Lactococcus petauri, a new species, has recently been implicated as another etiological agent. Both species cannot be distinguished by routine methods commonly used in diagnostic laboratories, resulting in their misidentification. In the present study, the identification of 48 isolates initially identified as L. garvieae was studied by determining their in-silico DNA–DNA hybridization (dDDH) and average nucleotide identity (ANI) values using pairwise comparisons of their whole genome sequences and the genomes of the type strains of L. garvieae and L. petauri. The genome sequences of 37 isolates from countries in which lactococcosis can be considered endemic (Spain, Italy, Türkiye, and Greece) were obtained in this study, and the genomes of 11 isolates were retrieved from the GenBank database. Isolates from Italy, Singapore, Japan, South Korea, India, one Turkish isolate from 2013 and two Spanish isolates recovered in 1992 and 1996 were confirmed as L. garvieae. The remaining isolates from Spain and Türkiye, as well as those from Portugal, Israel, USA, and Greece were identified as L. petauri. Overall, 60.4% of isolates previously identified as L. garvieae were found to be L. petauri. These results confirm the implication of both species in the etiology of lactococcosis and suggest that L. petauri plays a significant role in the epidemiology of this disease. Some of the isolates identified as L. petauri in the present study were isolated three decades ago, indicating that its association with lactococcosis is older than might be expected from the recent descriptions. The commercial Rapid ID32 Strep system was unable to discriminate between L. garvieae and L. petauri. However, both species exhibited some biochemical differences that might serve as phenotypic markers for their presumptive recognition. Consequently, isolates that hydrolyze hippurate and produce acid from sucrose and tagatose could be presumptively recognized as L. petauri, while those that fail these tests could be identified as L. garvieae. The results of this work indicate that great attention should be given to L. petauri in the epidemiology of lactococcosis.

Infective endocarditis following urinary tract infection caused by Globicatella sanguinis

We report the first case of infective endocarditis following urinary tract infection (UTI) caused by Globicatella sanguinis in an 87-year-old Japanese woman with recurrent episodes of UTI. We identified the pathogen using the Rapid ID32 Strep system. Accurate identification of this infection is important and essential for the effective antimicrobial coverage to this pathogen.

Enterococcus crotali sp. nov., isolated from faecal material of a timber rattlesnake.

A facultatively anaerobic, Gram-stain-positive bacterium, designated ETRF1T, was found in faecal material of a timber rattlesnake (Crotalus horridus). Based on a comparative 16S rRNA gene sequence analysis, the isolate was assigned to the genus Enterococcus. The 16S rRNA gene sequence of strain ETRF1T showed >97 % similarity to that of the type strains of Enterococcus rotai, E. caccae, E. silesiacus, E haemoperoxidus, E. ureasiticus, E. moraviensis, E. plantarum, E. quebecensis, E. ureilyticus, E. termitis, E. rivorum and E. faecalis. The organism could be distinguished from these 12 phylogenetically related enterococci using conventional biochemical testing, the Rapid ID32 Strep system, comparative pheS and rpoA gene sequence analysis, and comparative whole genome sequence analysis. The estimated in silico DNA-DNA hybridization values were <70 %, and average nucleotide identity values were <96 %, when compared to these 12 species, further validating that ETRF1T represents a unique species within the genus Enterococcus. On the basis of these analyses, strain ETRF1T (=CCUG 65857T=LMG 28312T) is proposed as the type strain of a novel species, Enterococcus crotali sp. nov.

Antimicrobial susceptibility of viridans group streptococcal blood isolates to eight antimicrobial agents.

A total of 155 viridans group streptococci blood culture isolates identified by the Rapid ID32 Strep system were tested for their minimum inhibitory concentrations (MICs) to penicillin, amoxicillin, ceftriaxone, erythromycin, clindamycin, rifampicin, vancomycin and teicoplanin using the E-test. The following species were identified: S. oralis (n = 67), S. mitis (n = 66), S. sanguis (n = 7), S. salivarius (n = 5), S. parasanguis (n = 4), S. gordonii (n = 3) and S. mutans (n = 3). S. oralis and S. mitis demonstrated the highest levels of resistance to the agents tested. There were 27% of S. oralis isolates resistant to pencillin, 51% resistant to erythromycin and 6% resistant to clindamycin. For S. mitis 11% were resistant to penicillin, 40% resistant to erythromycin and 3% resistant to clindamycin. Penicillin resistant isolates (MIC > or = 2 mg/l) also demonstrated decreased susceptibility to other antimicrobial agents tested in this study. High level resistance (MIC > or = 2 mg/l) to ceftriaxone was found in 12 isolates. The isolates identified as ceftriaxone resistant comprised S. oralis (n = 7), S. mitis (n = 4) and S. parasanguis (n = 1). This study has highlighted the difference in susceptibility between different species of viridans group streptococci. These findings are of concern in the light of spread of antibiotic resistance genes from S. oralis and S. mitis to the more invasive pneumococcus.

Genetic heterogeneities and phenotypic characteristics of strains of the genus Abiotrophia and proposal of Abiotrophia para-adiacens sp. nov.

The genus Abiotrophia represents a heterogeneous group of fastidious cocci that show a dependence on pyridoxal hydrochloride analogs for growth. The genetic heterogeneity in the genus Abiotrophia was examined by DNA-DNA hybridization, PCR assay of genomic DNA sequences, and restriction fragment length polymorphism and sequence homology analyses of the PCR-amplified 16S rRNA gene. Nine type or reference strains of Abiotrophia defectiva, Abiotrophia adiacens, and Abiotrophia elegans and 36 oral Abiotrophia isolates including the ones presumptively identified as Gemella morbillorum by the rapid ID32 STREP system were divided into four groups: A. defectiva (genotype 1), A. adiacens (genotype 2), A. elegans (genotype 4), and a fourth species (genotype 3) which we propose be named Abiotrophia para-adiacens sp. nov. A PCR assay specific for detection and identification of the novel Abiotrophia species was developed. A. para-adiacens generally produced beta-glucosidase but did not produce alpha- or beta-galactosidase or arginine dihydrolase, did not ferment, trehalose, pullulan, or tagatose, and was serotype IV, V, or VI. Thus, it was distinguished phenotypically from A. adiacens, A. elegans, and A. defectiva as well as, apparently, from the recently described species Abiotrophia balaenopterae sp. nov., which produces arginine dihydrolase and which ferments pullulan but not sucrose (P. A. Lawson et al., Int. J. Syst. Bacteriol. 49:503-506, 1999). Strain ATCC 27527, currently listed as G. morbillorum, was a member of the species A. para-adiacens.

Isolation and properties of bacteriolytic enzyme-producing cocci from the human mouth

One-hundred-and-one bacteriolytic enzyme-producing organisms were isolated from various sites of the mouth. All were non-hemolytic, Gram-positive, and chain-forming cocci. Ninety-one strains, like the reference strains of Streptococcus defectivus and S. adjacens, were dependent on pyridoxal for growth and produced a chromophore. The Rapid ID32 STREP system speciated these isolates as S. defectivus, S. adjacens or Gemella morbillorum. The remaining 10 bacteriolytic isolates were pyridoxal-independent and 8 belonged to S. intermedius. Some pyridoxal-independent S. intermedius reference strains including ATCC27335 T and all group D Enterococcus strains tested were also bacteriolytic. Thus, bacteriolytic enzyme production is common to nutritionally variant streptococci but not unique to S. defectivus and S. adjacens. The nutritionally variant strains generally had arylamidases but not alkaline phosphatase. The S. defectivus strains produced α- and β-galactosidases (biotype 1) whereas the S. adjacens strains generally produced N-acetyl-β-glucosaminidase and some had β-glucuronidase but others did not (biotypes 2 and 3). The G. morbillorum strains had no detectable activity of these glycosidases (biotype 4) but produced a chromophore and an arginine dihydrolase, exhibiting a physiological profile atypical of the Gemella species. This indicates the possible presence of an additional phenotypic group or a new species among the nutritionally variant streptococci.

Isolation and properties of bacteriolytic enzyme-producing cocci from the human mouth.

One-hundred-and-one bacteriolytic enzyme producing organisms were isolated from various sites of the mouth. All were non-hemolytic, Gram-positive, and chain-forming cocci. Ninety-one strains, like the reference strains of Streptococcus defectivus and S. adjacens, were dependent on pyridoxal for growth and produced a chromophore. The Rapid ID32 STREP system speciated these isolates as S. defectivus, S. adjacens or Gemella morbillorum. The remaining 10 bacteriolytic isolates were pyridoxal-independent and 8 belonged to S. intermediate. Some pyridoxal-independent S. intermedius reference strains including ATCC27335T and all group D Enterococcus strains tested were also bacteriolytic. Thus, bacteriolytic enzyme production is common to nutritionally variant streptococci but not unique to S. defectives and S. adjacens. The nutritionally variant strains generally had arylamidases but not alkaline phosphatase. The S. defectivus strains produced alpha-and beta-galactosidases (biotype 1) whereas the S. adjacens strains generally produced N-acetyl-beta-glucosaminidase and some had beta-glucuronidase but others did not (biotypes 2 and 3). The C. morbillorum strains had no detectable activity of these glycosidases (biotype 4) but produced a chromophore and an arginine dihydrolase, exhibiting a physiological profile atypical of the Gemella species. This indicates the possible presence of an additional phenotypic group or a new species among the nutritionally variant streptococci.

Comparative evaluation of Vitek Gram-positive identification system and API Rapid Strep system for identification of Streptococcu species of bovine origin

The Vitek Gram-positive identification system (GPI, Vitek Systems, Inc., Hazelwood, MO) and the API Rapid Strep system (Analytab Products, Plainview, NY) were evaluated for species identification of streptococci isolated from bovine mammary glands and compared to conventional biochemical methods. A total of 144 strains including Streptococcus uberis (60), S. dysgalactine (32), S. agalactiae (15), S. bovis (15), Enterococcus faecium (10) and Ent. faecalis (12) were evaluated. All references strains were identified correctly by both systems. Vitek GPI card system identified 94.4% of strains, including 95% od S. uberis, 93.8% of S. dysgalactiae, 93.3% of S. agalactiae and S. bovis II, 90% of Ent. faecium and 100% of Ent. faecalis. Majority of strains were identified with a 90–99% level of confidence, with an average of 8 h needed for identification. The API Rapis Strep system identified 96.5% of strains correctly, incluidng 95% of S. uberis, 96.6% of S. dysgalactiae, 93.3% of S. agalactiae, and 100% of S. bovis II, Ent. faecium, and Ent. faecalis. Majority pf strains were identified with excellent level of identification. With the exception of S. uberis, most strains were identified at 4 h of incubation.

Evaluation of the Rapid Strep System for Identification of Gram-Positive, Catalase-Negative Cocci Isolated from Bovine Intramammary Infections

The Rapid Strep system (Analytab Products, Plainview, NY) was used to identify 199 gram-positive, catalase-negative cocci isolated from bovine intramammary infections. The system accurately identified 88.4% of isolates. The system identified 100% of 46 Streptococcus agalactiae, 100% of 48 Streptococcus dysgalactiae, 54.5% of 11 Streptococcus equinus, and 96.2% of 53 Streptococcus uberis isolates. Enterococcus spp. were identified correctly 83.3% of the time. One of 4 Streptococcus saccharolyticus strains was identified as Streptococcus bovis, the previous classification for this organism, and 8 Streptococcus equi ssp. equi strains were misidentified as Streptococcus dysgalactiae. The Rapid Strep system was determined to be an acceptable alternative to conventional methods for identification of gram-positive, catalase-negative cocci isolated from bovine intramammary infections.

Physiological Characteristics and Deoxyribonucleic Acid Relatedness of Streptococcus intermedius Strains

Fifteen strains of Streptococcus intermedius and three mannitol-fermenting S. intermedius-like isolates from urine were studied by the S1 nuclease DNA homology assay and two physiological characterization methods. Three distinct homology groups (I, II, and III) were found. The three mannitol-fermenting S. intermedius-like strains and 2 of the 15 strains of S. intermedius did not fall within these groups. Each of the distinct homology groups could be distinguished by physiological testing. Homology group I strains were bile esculin positive (BE+), starch negative (Sta−), non-CO2 dependent, and melibiose negative (mel−); homology group II strains were BE−, Sta−, CO2 dependent, and mel−; homology group III strains were BE−, Sta+, non-CO2 dependent, and mel+. The two nonhomologous S. intermedius strains and three S. intermedius-like isolates from were phenotypically distinct from those in homology groups I through III. Characterization by the Rapid Strep system (Analytab Products, Plainview, N.Y.) identified homology groups I and II as either “S. milleri” I or II and homology group III strains as S. sanguis I or II. One nonhomologous S. intermedius strain was identified as S. sanguis I, and the other “S. milleri” I and the S. intermedius-like mannitol-fermenting strains were identified as S. milleri III by this method. We conclude that homology groups I, II, and III and the S. intermedius-like isolates from urine may represent separate taxa, but we do not make such a proposal on the basis of our limited data.

An Evaluation of the API-20 Strep System (Rapid-Strep System)

The API-20 strep (API System S.A., La-Balme-les-Grottes, France), which is identical to the Rapid-STREP system, marketed in the U.S. (API system S.A., Moutalieu-Vercieu, France), was compared with conventional biochemical and serological methods for the identification of streptococci. Ten strains of each of 27 species were tested. When the API-20 strep was read after 4 h incubation there was a 39.4% agreement with conventional methods. When the galleries were read after 24 h this figure rose to 78.8%. Of the remaining strains, 17% could not be identified because the profiles were not in the data base and 4.2% were identified differently from the identification obtained by conventional methods. The API-20 strep system was easy to perform and results were obtained in 24 h. The use of the system in combination with serological grouping reduces the misidentification of Streptococcus sanguis and Streptococcus salivarius, which can be differentiated by testing for levan and/or dextran production.

Evaluation of the Rapid Strep system for the identification of clinical isolates of Streptococcus species.

A total of 247 strains of streptococci isolated from humans were tested for identification in the Rapid Strep system. The identification rates and identification levels were different for each Streptococcus species. Our data indicate that the Rapid Strep system will identify nearly all the beta-hemolytic Streptococcus species if serological procedures are used in conjunction with the rapid physiological procedures. Of the group D streptococci, 98% of the enterococci and 95% of the non-enterococci were correctly identified. Of the commonly occurring viridans species, 85% were correctly identified, but only 10% of the less frequently occurring viridans species were identified. A total of 90% of the Streptococcus pneumoniae and 60% of the Aerococcus strains were correctly identified.

Evaluation of the rapid strep system for species identification of streptococci.

The Rapid Strep system (API System S.A., Montalieu-Vercieu, France) was evaluated, without additional tests, in the identification of 209 streptococci. Organisms included 59 beta-hemolytic, 36 group D, 24 Streptococcus pneumoniae, and 90 viridans group streptococci. The Rapid Strep system correctly identified to species level 69.5% of the beta-hemolytic strains, 100% of the group D strains, none of the S. pneumoniae strains, and 84.5% of the viridans group streptococci. The method provided excellent identification rates of groups A, B, and D but failed to differentiate between groups C and G. The method for preparation of suspensions for the Rapid Strep system, as initially recommended by the manufacturer, was responsible for the failure to identify S. pneumoniae. This method was subsequently modified to yield a heavier inoculum; all 10 pneumococcal strains tested with the revised inoculum method were correctly identified. Good identification rates of commonly encountered viridans strains were found. The Rapid Strep system represents a worthwhile advance in streptococcal species identification, especially for group D and viridans strains.

Use of the Rapid STREP system for identification of viridans streptococcal species.

A commercially available streptococcal identification system, Rapid STREP (API System S.A., Montalieu-Vercieu, France), was used to identify 119 viridans strains at the species level. A total of 92% of the strains tested were identified correctly with the Rapid STREP system, and 81% of the correct identifications were made at percentages of identification of 90 or greater. From these data it appears that the Rapid STREP system is a fairly accurate and rapid (24-h incubation) method for identifying species of viridans streptococci.