a 74-year - old man startled his dog and sustained a penetrating bite to his hand .
a few days later , the patient developed fever , chills and weakness , resulting in a fall .
significant comorbidities included chronic obstructive pulmonary disease , hypertension , dyslipidemia and idiopathic dilated cardiomyopathy , grade 3 left ventricle .
physical examination revealed a temperature of 40c , with bronchial breath sounds and crackles heard in the left chest ; the bite wound appeared improved .
blood cultures grew coagulase - negative staphylococcus , which was considered to be a contaminant , and gram - negative coccobacilli were later determined to be pasteurella dagmatis .
treatment was initiated with oral azithromycin 500 mg per day and intravenous ceftriaxone 1 g every 24 h for five days .
two sets of blood cultures were drawn 5 h arpart from the patient on the day of admission using bd bactec plus aerobic / f and anaerobic / f bottles ( becton , dickinson and company , canada ) .
three bottles produced gram - negative coccobacilli with beaded ends in 14 h to 34 h , and one anaerobic / f bottle also produced gram - positive cocci in clusters , which were subsequently identified as coagulase - negative staphylococci ; this organism was considered to be a contaminant . the subcultures on blood agar plates produced tiny grey - brown creamy colonies , which did not grow on macconkey agar .
the gram - negative coccobacilli were initially identified as pasturella pneumotropica by the vitek 2 system , software version 06.01 ( biomerieux , france ) using the gn card , with bionumber 0001010210040001 and an excellent identification ( probability 99% ) .
unusual bacteria such as this are routinely sent to the local reference laboratory ( public health ontario , toronto , ontario ) for confirmation of identification and susceptibility testing . the susceptibility profile of the bacterium was interpreted by clsi m45-a2 ( 1 ) ( table 1).the biochemical characteristics ( table 2 ) , 16s ribosomal rna ( rrna ) gene polymerase chain reaction ( pcr ) and sequencing ( below ) , as well as matrix - assisted laser desorption / ionization time - of - flight mass spectrometry ( maldi - tof ms ) were used for identification of the bacterium .
traditional biochemical testing was performed on the isolate and based on its profile was determined to be p dagmatis ( 2 ) ( table 2 ) . because this organism is not often encountered , alternate identification methods were also used to ensure a correct identification .
a 736-base pair amplicon was generated ( primers , forward : 5agtttgatcctggctcag3 ; reverse : 5 ggactaccagggtatctaat3 ) and sequenced using routine methods ( 3 ) .
the sequence was analyzed using national center for biotechnology information basic local alignment search tool ( 4 ) and results were interpreted using clsi mm18-a guidelines ( 5 ) .
the pcr product was 99% similar to six deposits within the nr / nt database with 99% to 100% coverage .
sequences with high levels of homology to the query sequence included the type strain of p dagmatis , atcc 43325/ccug 12397 ( 99% ; nr_042883.1 and m75051.1 ) and the type strain of pasteurella stomatis , ccug 17979 ( 99% ; nr_042888.1 ) .
based on clsi mm18-a interpretation guidelines ( 5 ) , due to the low level of demarcation of the sequence of the 16s rrna gene between these species , the unknown bacteria may only be identified as p dagmatis or p stomatis . however , based on the biochemical profile ( table 2 ) , this organism could not be p stomatis ( which is urease and maltose negative , because the organism in question is urease and maltose positive ) ; therefore , in the present case , the identification of the organism was p dagmatis .
two sets of blood cultures were drawn 5 h arpart from the patient on the day of admission using bd bactec plus aerobic / f and anaerobic / f bottles ( becton , dickinson and company , canada ) .
three bottles produced gram - negative coccobacilli with beaded ends in 14 h to 34 h , and one anaerobic / f bottle also produced gram - positive cocci in clusters , which were subsequently identified as coagulase - negative staphylococci ; this organism was considered to be a contaminant .
the subcultures on blood agar plates produced tiny grey - brown creamy colonies , which did not grow on macconkey agar .
the gram - negative coccobacilli were initially identified as pasturella pneumotropica by the vitek 2 system , software version 06.01 ( biomerieux , france ) using the gn card , with bionumber 0001010210040001 and an excellent identification ( probability 99% ) .
unusual bacteria such as this are routinely sent to the local reference laboratory ( public health ontario , toronto , ontario ) for confirmation of identification and susceptibility testing .
the susceptibility profile of the bacterium was interpreted by clsi m45-a2 ( 1 ) ( table 1).the biochemical characteristics ( table 2 ) , 16s ribosomal rna ( rrna ) gene polymerase chain reaction ( pcr ) and sequencing ( below ) , as well as matrix - assisted laser desorption / ionization time - of - flight mass spectrometry ( maldi - tof ms ) were used for identification of the bacterium .
traditional biochemical testing was performed on the isolate and based on its profile was determined to be p dagmatis ( 2 ) ( table 2 ) . because this organism is not often encountered , alternate identification methods were also used to ensure a correct identification .
a 736-base pair amplicon was generated ( primers , forward : 5agtttgatcctggctcag3 ; reverse : 5 ggactaccagggtatctaat3 ) and sequenced using routine methods ( 3 ) .
the sequence was analyzed using national center for biotechnology information basic local alignment search tool ( 4 ) and results were interpreted using clsi mm18-a guidelines ( 5 ) .
the pcr product was 99% similar to six deposits within the nr / nt database with 99% to 100% coverage .
sequences with high levels of homology to the query sequence included the type strain of p dagmatis , atcc 43325/ccug 12397 ( 99% ; nr_042883.1 and m75051.1 ) and the type strain of pasteurella stomatis , ccug 17979 ( 99% ; nr_042888.1 ) .
based on clsi mm18-a interpretation guidelines ( 5 ) , due to the low level of demarcation of the sequence of the 16s rrna gene between these species , the unknown bacteria may only be identified as p dagmatis or p stomatis .
however , based on the biochemical profile ( table 2 ) , this organism could not be p stomatis ( which is urease and maltose negative , because the organism in question is urease and maltose positive ) ; therefore , in the present case , the identification of the organism was p dagmatis .
single colonies of fresh organisms grown overnight were prepared using a modified formic acid extraction procedure and analyzed using the bruker maldi biotyper ( bruker daltonics , germany ) in duplicate using standard settings .
the query spectra had a high level of similarity > 2.0 ( 2.0 is an acceptable score for species - level identification ) to p dagmatis spectra within the routine commercial database .
the top five matches were to spectra from different strains of p dagmatis within the commercial database .
it is a gram - negative coccobacillus belonging to the pasteurellaceae family , which is fermentative , aerobic , nonmotile , oxidase positive and penicillin - sensitive .
this organism has been isolated from both dogs and cats as normal flora , and also reported as a pathogen in human infections .
it was previously labeled as pasteurella  gas  , pasteurella new species 1 or p pneumotropica type henriksen , and was eventually reclassified as p dagmatis ( 6 ) .
bacteria from the pasteurellaceae family cause zoonotic infections in humans , with p multocida and p canis being the most common pasturella species reported in human infections ( 7,8 ) .
infections caused by pasteurella species are typically introduced by animals , particularly cat or dog bites , but also occasionally by other animals , and often manifest as skin or soft tissue infections ( 79 ) .
sometimes , animal contact is not prominent in the initial patient history ( 10,11 ) ( table 3 ) .
the most probable route of transmission of p dagmatis infection in the present case was most likely the bite and licking of the patient s traumatized skin by his dog , as has been previously described ( 11,12 ) .
continuous shedding of p dagmatis from asymptomatic animals ( eg , in dog urine ) and whether it can be an indirect route of infection to human remains to be investigated . while pasteurella species are well recognized for causing skin or soft tissue infections , p dagmatis can also cause more serious disease , including infective and prosthetic valve endocarditis ( 10,14,15 ) , septicemia ( 11,12,16 ) , peritonitis ( 17 ) , vertebral osteomyelitis ( 18,19 ) , chronic bronchiectasis ( 20 ) and pneumonia ( 21 ) , mainly in immunocompromised patients .
a small number of case reports describing systemic human p dagmatis infections are listed in table 3 .
interestingly , while pasturella species infrequently cause systemic infectious disease , in our review of the literature , when p dagmatis infections are reported , they appear to be frequently associated with severe disseminated infection including bacteremia .
coinfections of p dagmatis with another pasturella species have also been observed ( 9,12,22 ) ; therefore , it is important for the laboratory to test multiple morphotypes from the plate to ensure that > 1 pasturella species is not present .
similar to other pasturella species , p dagmatis is typically highly susceptible to many antibiotics , particularly , the beta - lactams ( table 1 ) .
early suspicion and timely laboratory diagnosis of pasturella infection are crucial for a favourable clinical outcome .
several reports have demonstrated that the vitek 2 gn card misidentifies p dagmatis as p pneumotropica or p canis , despite an excellent identification probability ( 9,15,23,24 ) .
this is most likely because p dagmatis has not been included in the system database ; as well , there has been a nomenclature change because p dagmatis was formerly grouped with p pneumotropica , type henriksen . in a study that included 66
clinical pasturella isolates and used soda gene sequencing as a reference method , zangenah et al ( 24 ) revealed that vitek 2 only identified approximately 50% of pasturella isolates correctly , while conventional biochemical tests and maldi - tof ms were able to correctly identify 94% and 89% , respectively .
interestingly , in the zangenah et al ( 24 ) study , two p dagmatis isolates were not identified by vitek ms maldi - tof ( biomerieux , france ) and this limitation was also observed in our study ( data not shown ) . the biological and genetic profiles among p dagmatis , p pneumotropica and p stomatis are very similar ( table 2 ) ; both a commercial biochemical identification system and the sequence analysis of a portion of the 16s rrna gene were unable to differentiate between these species .
correct identification was made using maldi - tof ms ( maldi biotyper , bruker , germany ) and was also supported by comparing the key biochemical characteristics among p dagmatis , p pneumotropica and p stomatis ( table 2 ) .
it is probable that many clinical isolates of p dagmatis have been misidentified due to the limitation of commercial biochemical identification systems , such as vitek 2 .
misidentification may have contributed to an underestimation of the frequency of this organism in clinical samples ; however , the growing use of maldi - tof ms systems for microorganism identification in routine clinical microbiology laboratories may allow for a more accurate picture of how frequently p dagmatis causes infections .
correct identification is important for diagnosis and therapeutic management , and epidemiological monitoring of the transmission of pasturella species , particularly for the systemic infections such as in the present case .
unfortunately , most routine methods available at hospital laboratories can not identify the organism correctly .
p dagmatis can cause severe animal - associated infections in humans , mainly in immunocompromised individuals . to our knowledge , this is the first systemic p dagmatis infection reported in canada .
clinical outcomes rely on early accurate laboratory confirmation and timely administration of effective antibiotic treatment . conventional identification of p dagmatis using vitek 2 can be misleading , probably due to the absence of this organism from the database ; 16s rrna gene sequence analysis and maldi - tof ms systems represent excellent options for identifying rarely encountered or difficult to identify organisms , such as members of the pasturellaceae family .
the present study re - emphasizes the need for continuously improving the database of automatic microbial identification systems .
 pasteurella dagmatis , a gram - negative coccobacillus , has been isolated from both dogs and cats as normal flora . 
 it is also a fairly new species for many clinicians because it is a pathogen in human infections . 
 the authors present a case of bacteremia in a 74-year - old man that was caused by p dagmatis . 
 a comparison of other reported cases of bacteremia due to p dagmatis is provided , along with a discussion of the challenges of standard automatic identification including alternative methodologies . 