Isolation of Potentially Novel Brucella spp. from Frogs (original) (raw)
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Journal of Zoo and Wildlife Medicine
Two novel and distinct Brucella strains were recovered from 5 of 10 adult, sex undetermined, captive waxy tree frogs (Phyllomedusa sauvagii) and two of five adult, sex undetermined, captive Colorado river toads (Incilius alvarius) held in a zoologic collection with clinical and pathologic findings of bacterial disease. These amphibians originated from three separate private breeding facilities over several years and exhibited disease 9-49 mo following release from quarantine. Common presenting signs were vague but included focal abscessation, weight loss, change in coloration, anorexia, and decreased perching. Two waxy tree frogs and one Colorado river toad recovered with supportive care and antimicrobial treatment based on susceptibility testing. Microgranulomatosis, subcutaneous and renal abscessation, femoral osteomyelitis, and multicentric infection were the most common histologic findings. The organisms were identified antemortem in samples from subcutaneous abscesses, cloaca, and skin and from a variety of organ systems postmortem, and demonstrated a consistent susceptibility pattern. Initial isolates were misidentified as Ochrobactrum anthropi. Polymerase chain reaction and sequencing of the 16S rRNA gene identified the two organisms as novel Brucella strains similar to Brucella inopinata-like sp. and other novel organisms within the emerging ''BO clade.'' Brucella strain oaks (isolated from waxy tree frogs) and Brucella strain leathers (isolated from Colorado river toads) differed from each other by 16 of 571 base pairs in a region of chromosome 2, and did not closely match any previous GenBank entries. This report describes the clinicopathologic features of infection by these bacteria in two amphibian species and expands the range of novel Brucella organisms from amphibian reservoirs.
Frontiers in Veterinary Science, 2018
Several Brucella isolates have been described in wild-caught and "exotic" amphibians from various continents and identified as B. inopinata-like strains. On the basis of epidemiological investigations conducted in June 2017 in France in a farm producing domestic frogs (Pelophylax ridibundus) for human consumption of frog's legs, potentially pathogenic bacteria were isolated from adults showing lesions (joint and subcutaneous abscesses). The bacteria were initially misidentified as Ochrobactrum anthropi using a commercial identification system, prior to being identified as Brucella spp. by MALDI-TOF assay. Classical phenotypic identification confirmed the Brucella genus, but did not make it possible to conclude unequivocally on species determination. Conventional and innovative bacteriological and molecular methods concluded that the investigated strain was very close to B. microti species, and not B. inopinata-like strains, as expected. The methods included growth kinetic, antimicrobial susceptibility testing, RT-PCR, Bruce-Ladder, Suis-Ladder, RFLP-PCR, AMOS-ERY, MLVA-16, the ectoine system, 16S rRNA and recA sequence analyses, the LPS pattern, in silico MLST-21, comparative whole-genome analyses (including average nucleotide identity ANI and whole-genome SNP analysis) and HRM-PCR assays. Minor polyphasic discrepancies, especially phage lysis and A-dominant agglutination patterns, as well as, small molecular divergences suggest the investigated strain should be considered a B. microti-like strain, raising concerns about its environmental persistence and unknown animal pathogenic and zoonotic potential as for other B. microti strains described to date.
PloS one, 2016
The genus Brucella comprises various species of both veterinary and human medical importance. All species are genetically highly related to each other, sharing intra-species average nucleotide identities (ANI) of > 99%. Infections occur among various warm-blooded animal species, marine mammals, and humans. Until recently, amphibians had not been recognized as a host for Brucella. In this study, however, we show that novel Brucella species are distributed among exotic frogs worldwide. Comparative recA gene analysis of 36 frog isolates from various continents and different frog species revealed an unexpected high genetic diversity, not observed among classical Brucella species. In phylogenetic reconstructions the isolates consequently formed various clusters and grouped together with atypical more distantly related brucellae, like B. inopinata, strain BO2, and Australian isolates from rodents, some of which were isolated as human pathogens. Of one frog isolate (10RB9215) the genome...
Brucella microti‐ like prevalence in French farms producing frogs
Transboundary and Emerging Diseases, 2019
Brucellosis affects a wide range of animal hosts, generally causing reproductive disorders resulting in significant economic losses in farmed species. It is recognized as one of the most important bacterial zoonoses worldwide with 500,000 human cases per year (Godfroid et al., 2005). The genus Brucella comprises twelve species historically differentiated by host specificity and phenotypic characteristics (OIE, 2016). Brucella spp. have been evidenced in various hosts belonging to different ecosystems. Beside the six classical species (B. abortus, B melitensis, B. suis, B. canis, B. neotomae and B. ovis) affecting terrestrial mammals (OIE, 2016), novel members have been lately added to this group (B. pinnipedialis, B.ceti and B. papionis), thus extending the variety of Brucella animal hosts to marine mammals and primates (Foster, Osterman, Godfroid, Jacques, & Cloeckaert, 2007; Whatmore et al., 2014). Three other novel species genetically related to an atypical group of the genus (Scholz & Vergnaud, 2013) were first isolated in warm-blooded hosts: B. microti in common voles and foxes (
Scientific reports, 2017
Twenty-one small Gram-negative motile coccobacilli were isolated from 15 systemically diseased African bullfrogs (Pyxicephalus edulis), and were initially identified as Ochrobactrum anthropi by standard microbiological identification systems. Phylogenetic reconstructions using combined molecular analyses and comparative whole genome analysis of the most diverse of the bullfrog strains verified affiliation with the genus Brucella and placed the isolates in a cluster containing B. inopinata and the other non-classical Brucella species but also revealed significant genetic differences within the group. Four representative but molecularly and phenotypically diverse strains were used for in vitro and in vivo infection experiments. All readily multiplied in macrophage-like murine J774-cells, and their overall intramacrophagic growth rate was comparable to that of B. inopinata BO1 and slightly higher than that of B. microti CCM 4915. In the BALB/c murine model of infection these strains re...
Isolation of Brucella from a White’s tree frog (Litoria caerulea)
JMM Case Reports, 2015
Introduction: Brucellosis is a zoonotic disease that has a significant economic, social and public health impact in many parts of the world. The causative agents are members of the genus Brucella currently comprising 11 species and with an expanding known host range in recent years. Case presentation: One of a pair of White's tree frogs (Litoria caerulea) developed skin lesions from which a pure growth of a haemolytic organism was obtained. The isolate was identified as Brucella melitensis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, although the colony morphology was inconsistent with this identification. Applying the classical biotyping approach used to subdivide members of the genus Brucella, the isolate did not correspond to any known Brucella sp. However, PCR targeting of genes specific for members of the genus Brucella was strongly positive and 16S rRNA gene sequencing revealed a close relationship with extant Brucella spp. In order to place the isolate more accurately, a multilocus sequencing approach was applied, which confirmed that the isolate represented a novel member of the emerging 'atypical' Brucella group, which includes isolates from human disease, from rodents and, more recently, reported isolations from frogs in Germany. Conclusion: This case represents the first report of isolation of a Brucella sp. from frogs outside Germany and suggests that these isolates may be widespread. Whilst there is no evidence to date that these isolates represent a zoonotic threat, the association of other 'atypical' Brucella sp. with human disease suggests that appropriate measures should be taken to avoid unnecessary contact with potentially infected amphibians until the zoonotic potential of this group is better understood.
Brazilian Journal of Animal and Environmental Research, 2021
Ambystoma velasci, is a natural inhabitant of the crater lakes Atexcac, La Preciosa and Quechulac, in the Eastern Basin, Puebla, Mexico. Currently, their populations are at risk from the loss and microbiological contamination of their habitat. Brucella spp., is a zoonotic bacterium, which has been expanding to new taxa as reservoirs. The objective of this project was to detect and isolate Brucella sp., in Ambystoma velasci as a new host. Between February 2014 and April 2017, 25 specimens of A. velasci were collected. The primo-isolation were recovered from samples of the digestive, reproductive and excretory system (cloaca), and identified by standard microbiological and molecular tests, obtaining (40%) of positive results. They were compared by PCR, with two reference vaccine strains (BM16 and BS19), the bp 26 (IS711) gene was amplified, which is specific
Journal of Medical Microbiology, 2008
Ochrobactrum anthropi, Ochrobactrum intermedium and Brucella spp. are phenotypically and genetically closely related pathogens that may cause disease with similar clinical presentation. Consequently, difficulties in their identification and differentiation have been reported. In this study, a sensitive recA gene-based multi-primer single-target PCR (MP-ST-PCR) was developed that allowed the specific detection and differentiation of these clinically relevant pathogens. The specificity of the assay was evaluated using a representative panel of 50 O. anthropi and 16 O. intermedium strains and the type strains of all Brucella spp. Detection limits for purified DNA from O. anthropi, O. intermedium and Brucella melitensis were 100, 10 and 100 fg, respectively. Brucella DNA was also successfully detected in various clinical specimens from a human patient with culture-proven brucellosis and from a Brucella-infected sheep and its aborted fetuses. The sensitivity of the MP-ST-PCR was comparable to that of an evaluated in-house Brucella real-time PCR assay. The developed assay closes a diagnostic gap and provides a simple but robust tool for the sensitive detection and correct identification of O. anthropi, O. intermedium and Brucella spp.
Brucella suis bacteremia misidentified as Ochrobactrum anthropi by the VITEK 2 system
The Journal of Infection in Developing Countries, 2016
Ochrobactrum and Brucella are genetically related genera of the family Brucellaceae, sharing 98.8% rRNA similarity. Because of their phenotypic similarity, Ochrobactrum can be miscoded as Brucella by automated identification systems. The misidentification on blood cultures (BCs) of B. suis as O. anthropi by the VITEK 2 system is herein described. A 67-year-old male with a prosthetic mitral valve and fever was admitted with bacteremia due to a Gram-negative coccobacillus identified as O. anthropi by VITEK 2. The patient’s fever persisted along with positive blood cultures despite specific antimicrobial treatment. Due to this adverse outcome, the patient was interrogated again and admitted having domestic swine. Serological tests were positive for acute brucellosis. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) of BC strains identified B. suis biovar 1. Timely identification of Brucella is essential for providing proper treatment to the patient and for ...