Aptamer selection against cell extracts containing the zoonotic obligate intracellular bacterium, Anaplasma phagocytophilum.

A. phagocytophilum is a zoonotic and tick-borne bacterium, threatening human and animal health. Many questions persist concerning the variability of strains and the mechanisms governing the interactions with its different hosts. These gaps can be explained by the difficulty to cultivate and study A. phagocytophilum because of its strict intracellular location and the lack of specific tools, in particular monoclonal antibodies, currently unavailable. The objective of our study was to develop DNA aptamers against A. phagocytophilum, or molecules expressed during the infection, as new study and/or capture tools. Selecting aptamers was a major challenge due to the strict intracellular location of the bacterium. To meet this challenge, we set up a customized selection protocol against an enriched suspension of A. phagocytophilum NY18 strain, cultivated in HL-60 cells. The implementation of SELEX allowed the selection of three aptamers, characterized by a high affinity for HL-60 cells infected with A. phagocytophilum NY18 strain. Interestingly, the targets of these three aptamers are most likely proteins expressed at different times of infection. The selected aptamers could contribute to increase our understanding of the interactions between A. phagocytophilum and its hosts, as well as permit the development of new diagnostic, therapeutic or drug delivery appliances.

Genomic Characterization of Three Novel Bartonella Strains in a Rodent and Two Bat Species from Mexico.

Rodents and bats are the most diverse mammal group that host Bartonella species. In the Americas, they were described as harboring Bartonella species; however, they were mostly characterized to the genotypic level. We describe here Bartonella isolates obtained from blood samples of one rodent (Peromyscus yucatanicus from San José Pibtuch, Yucatan) and two bat species (Desmodus rotundus from Progreso, and Pteronotus parnellii from Chamela-Cuitzmala) from Mexico. We sequenced and described the genomic features of three Bartonella strains and performed phylogenomic and pangenome analyses to decipher their phylogenetic relationships. The mouse-associated genome was closely related to Bartonella vinsonii. The two bat-associated genomes clustered into a single distinct clade in between lineages 3 and 4, suggesting to be an ancestor of the rodent-associated Bartonella clade (lineage 4). These three genomes showed <95% OrthoANI values compared to any other Bartonella genome, and therefore should be considered as novel species. In addition, our analyses suggest that the B. vinsonii complex should be revised, and all B. vinsonii subspecies need to be renamed and considered as full species. The phylogenomic clustering of the bat-associated Bartonella strains and their virulence factor profile (lack of the Vbh/TraG conjugation system remains of the T4SS) suggest that it should be considered as a new lineage clade (L5) within the Bartonella genus.

Optimized Lambda Exonuclease Digestion or Purification Using Streptavidin-Coated Beads: Which One Is Best for Successful DNA Aptamer Selection?

The high failure rate of the in vitro aptamer selection process by SELEX (Systematic Evolution of Ligands by EXponential enrichment) limits the production of these innovative oligonucleotides and, consequently, limits their potential applications. The generation of single-stranded DNA (ssDNA) is a critical step of SELEX, directly affecting the enrichment and the selection of potential binding sequences. The main goal of this study was to confirm the best method for generating ssDNA by comparing the purification of ssDNA, using streptavidin-coated beads, and lambda exonuclease digestion, and by improving ssDNA recovery through protocol improvements. In addition, three techniques for quantifying the ssDNA generated (Qubit vs. NanodropTM vs. gel quantification) were compared, and these demonstrated the accuracy of the gel-based quantification method. Lambda exonuclease digestion was found to be more efficient for ssDNA recovery than purification using streptavidin-coated beads, both quantitatively and qualitatively. In conclusion, this work provides a detailed and rigorous protocol for generating ssDNA, improving the chances of a successful aptamer selection process.

Enterobacterales high-risk clones and plasmids spreading blaESBL/AmpC and blaOXA-48 genes within and between hospitalized dogs and their environment.

BACKGROUND: Compared with healthcare settings, the role of veterinary hospitals in the spread of extended-spectrum cephalosporin- and carbapenem-resistant (ESC-R/CP-R) bacteria has been overlooked. OBJECTIVES: To investigate using genome-based approaches the dynamics of ESC-R and CP-R Enterobacterales among 125 dogs admitted to the same veterinary hospital over a 4 month period. METHODS: Dogs (n = 125) were sampled within 48 h of admission and at discharge. ESC-R/CP-R were phenotypically characterized and whole-genome sequenced using short- and long-read technologies. Phylogenetic analyses were performed using appropriate pipelines. RESULTS: ESC-R/CP-R prevalence in dogs was 4.8% (6/125) upon admission and reached 24.8% (31/125) at discharge, reflecting multiple acquisitions of ESBL/AmpC and OXA-48-positive Enterobacterales during hospitalization. Indistinguishable or closely related isolates were found within dogs, shared between dogs, and shared between dogs and their environment, suggesting numerous clonal and plasmid spreads. Even though carbapenems are not licensed for use in companion animals, a wide distribution of the blaOXA-48/IncL plasmid was evidenced across different bacterial species and dogs. CONCLUSIONS: This study highlights nosocomial acquisitions of ESBL/AmpC and carbapenemase-producing Enterobacterales by companion animals and the risk of further transmission within the community in a One Health perspective. Reinforced infection prevention and control measures and screening procedures are urgently needed in small animal veterinary settings where advanced therapeutics and intensive care is provided.

Sensitivity of bovine tuberculosis surveillance through intradermal tests in cattle in France: An evaluation of different scenarios.

The current situation regarding bovine tuberculosis (bTB) in Europe is spatially heterogeneous, with stagnating or increasing trends in bTB prevalence in many European regions, underlying the challenge in controlling this disease. In France, in spite of the implementation of two control programs in 2010-2012 to eradicate the disease and maintain the bTB-free status, bTB prevalence has continued to increase, underlying the need to reinforce and adapt surveillance measures. The goal of this study was to evaluate the effectiveness of bTB surveillance in high-risk areas in metropolitan France, with an emphasis on the criteria to select herds and animals within herds in the context of programmed surveillance and movement testing. The fraction of bTB-infected herds detected by the surveillance was quantified using a stochastic scenario tree modelling approach, with input parameter values based on surveillance and cattle traceability data and literature. The detection fraction was assessed for the current surveillance system and for alternative scenarios. The model predicted that the median detection fraction of infected herds by the current programmed surveillance in high-risk areas, which consists in annual testing of herds with a minimum age of testing of 24 months, was 71.5 % (interquartile interval: 47.4-89.4). The results showed a significant gain of the detection fraction with a decrease from 24 to 12 months old (83.5 % [60.6-95.9]) or to six weeks old (91.3 % [71.6-99.0]). Regarding pre-movement surveillance, tests are currently mandatory for bovines that originate from a previously infected herd or from a herd epidemiologically linked to a bTB-infected herd. The median detection fraction predicted by the model for this surveillance scenario was 1.2 % [0.7-1.8]. For the alternative scenario, where surveillance would be extended to all herds in high-risk areas, the model predicted a significant increase of the detection fraction to 26.5 % [18.1-37.9]. The results were sensitive to the following input values: the number of infected bovines within herds and, to a lower extent, the comparative intradermal tuberculin test sensitivity for both models, and surveillance coverage for the model on pre-movement surveillance. Our study underlines several complementary ways to improve the detection of infected herds, which is critical for implementing control measures and epidemiological investigations as early as possible. These necessary changes in surveillance must be accompanied by a global reflexion on surveillance financing.

High-Throughput Microfluidic Real-Time PCR for the Detection of Multiple Microorganisms in Ixodid Cattle Ticks in Northeast Algeria.

Ixodid ticks are hematophagous arthropods considered to be prominent ectoparasite vectors that have a negative impact on cattle, either through direct injury or via the transmission of several pathogens. In this study, we investigated the molecular infection rates of numerous tick-borne pathogens in ticks sampled on cattle from the Kabylia region, northeastern Algeria, using a high-throughput microfluidic real-time PCR system. A total of 235 ticks belonging to seven species of the genera Rhipicephalus, Hyalomma, and Ixodes were sampled on cattle and then screened for the presence of 36 different species of bacteria and protozoans. The most prevalent tick-borne microorganisms were Rickettsia spp. at 79.1%, followed by Francisella-like endosymbionts (62.9%), Theileria spp. (17.8%), Anaplasma spp. (14.4%), Bartonella spp. (6.8%), Borrelia spp. (6.8%), and Babesia spp. (2.5%). Among the 80.4% of ticks bearing microorganisms, 20%, 36.6%, 21.7%, and 2.1% were positive for one, two, three, and four different microorganisms, respectively. Rickettsia aeschlimannii was detected in Hyalomma marginatum, Hyalomma detritum, and Rhipicephalus bursa ticks. Rickettsia massiliae was found in Rhipicephalus sanguineus, and Rickettsiamonacensis and Rickettsia helvetica were detected in Ixodesricinus. Anaplasma marginale was found in all identified tick genera, but Anaplasma centrale was detected exclusively in Rhipicephalus spp. ticks. The DNA of Borrelia spp. and Bartonella spp. was identified in several tick species. Theileria orientalis was found in R. bursa, R. sanguineus, H. detritum, H. marginatum, and I. ricinus and Babesia bigemina was found in Rhipicephalus annulatus and R. sanguineus. Our study highlights the importance of tick-borne pathogens in cattle in Algeria.

Enlisting the Ixodes scapularis Embryonic ISE6 Cell Line to Investigate the Neuronal Basis of Tick-Pathogen Interactions.

Neuropeptides are small signaling molecules expressed in the tick central nervous system, i.e., the synganglion. The neuronal-like Ixodes scapularis embryonic cell line, ISE6, is an effective tool frequently used for examining tick-pathogen interactions. We detected 37 neuropeptide transcripts in the I. scapularis ISE6 cell line using in silico methods, and six of these neuropeptide genes were used for experimental validation. Among these six neuropeptide genes, the tachykinin-related peptide (TRP) of ISE6 cells varied in transcript expression depending on the infection strain of the tick-borne pathogen, Anaplasma phagocytophilum. The immunocytochemistry of TRP revealed cytoplasmic expression in a prominent ISE6 cell subpopulation. The presence of TRP was also confirmed in A. phagocytophilum-infected ISE6 cells. The in situ hybridization and immunohistochemistry of TRP of I. scapularis synganglion revealed expression in distinct neuronal cells. In addition, TRP immunoreaction was detected in axons exiting the synganglion via peripheral nerves as well as in hemal nerve-associated lateral segmental organs. The characterization of a complete Ixodes neuropeptidome in ISE6 cells may serve as an effective in vitro tool to study how tick-borne pathogens interact with synganglion components that are vital to tick physiology. Therefore, our current study is a potential stepping stone for in vivo experiments to further examine the neuronal basis of tick-pathogen interactions.

Bovines Harbor a Diverse Array of Vector-Borne Pathogens in Northeast Algeria.

Arthropod-borne hemoparasites represent a serious health problem in livestock, causing significant production losses. Currently, the evidence of Anaplasma spp., Theileria spp., Babesia spp., and hemotropic Mycoplasma spp. in Algeria remains limited to a few scattered geographical regions. In this work, our objectives were to study the prevalence of these vector-borne pathogens and to search other agents not yet described in Algeria as well as the identification of statistical associations with various risk factors in cattle in the northeast of Algeria. Among the 205 cattle blood samples tested by PCR analysis, 42.4% positive results were obtained for at least one pathogen. The overall rates of Anaplasma spp., Theileria/Babesia spp., and Mycoplasma spp. in the cattle sampled were respectively 30.7%, 18.5%, and 2.9%; co-infections with multiple species was also detected. Anaplasma spp. and Theileria/Babesia spp. were detected at a higher rate in cattle under 3 years old, according to univariate analysis. Anaplasma spp. DNA was detected more frequently in our sample in cattle living in semi extensive farming. Our study provides additional data about Anaplasma spp., Theileria/Babesia spp. and reveals for the first time that Mycoplasma wenyonii and 'Candidatus Mycoplasma hemobos are present in cattle in Northeast Algeria.

First report on Bartonella henselae in dromedary camels (Camelus dromedarius).

Bartonellosis is one of the clinically underdiagnosed emerging bacterial diseases among domestic livestock, particularly in camels. Until now, the natural infection of camels with Bartonella species was not investigated in Tunisia. In the attempt of filling this gap in knowledge, a total of 412 dromedary camels (Camelus dromedarius) as well as 300 associated ticks (Hyalomma dromedarii (160; 53.4%), H. impeltatum (131; 43.6%) and H. excavatum (9; 3%) were screened for the presence of Bartonella spp. by PCR followed by a sequencing step through the amplification of the rpoB gene. Positive samples were then tested and further characterized by the combined use of the ftsZ and gltA genes. Fifteen camels (3.6%) were found to be positive to Bartonella spp. However, there was no evidence of Bartonella DNA in any of the analyzed ticks. Risk factors' analysis shows that camels derived from arid and semi-arid bioclimatic areas were more infected than those originated from desert area. Molecular characterization and phylogenetic analysis revealed the occurrence of novel B. henselae genotypes closely related to those isolated from humans, cats, and lions. By combining the characteristics of each single gene with those of concatenated sequences, we report here the first molecular detection of B. henselae in the dromedary camel suggesting a possible involvement of camelids as hosts or reservoirs in the transmission cycle of this emerging bacterium in arid and saharan areas.

Multiple locus variable number tandem repeat analysis for the characterization of wild feline Bartonella species and subspecies.

Bartonella genus includes an increasing number of species and subspecies, especially among wild felids, the positioning of which, with regards to the zoonotic species Bartonella henselae, is important to determine. The aim of this study was to test the ability of a molecular typing technique to distinguish between various Bartonella isolates obtained from four different species of free-ranging and captive wild felids and to identify key profiles or markers allowing differentiating them from each other and/or from B. henselae or B. koehlerae. A molecular typing technique for B. henselae based on the polymorphism of variable number tandem repeat units (VNTR) called MLVA (Multiple Locus VNTR Analysis) was applied to 24 Bartonella isolates from free-ranging or captive wild felids, 19 of which were obtained from California and five from three countries in Southern Africa, and compared with 49 B. henselae isolates from cats, dog or humans from the United States including the human ATCC (American Type Culture Collection) reference strain, B. henselae Houston 1. MLVA allowed distinguishing Bartonella isolates from wild felids from either B. henselae or B. koehlerae. We confirmed infection of semi-captive cheetahs with an isolate similar to a Californian bobcat isolate. MLVA also confirmed the unique profile of a free-ranging cheetah isolate from Namibia. Specific profiles were observed making MVLA a useful identification/classification tool of these wild felid isolates and suggesting that they are highly adapted to a specific feline reservoir. Finally, circulation of B. henselae isolates between domestic cats, wild felids and humans is likely occurring, based on the close allelic profiles of some isolates.

Bovine Organospecific Microvascular Endothelial Cell Lines as New and Relevant In Vitro Models to Study Viral Infections.

Microvascular endothelial cells constitute potential targets for exogenous microorganisms, in particular for vector-borne pathogens. Their phenotypic and functional variations according to the organs they are coming from provide an explanation of the organ selectivity expressed in vivo by pathogens. In order to make available relevant tools for in vitro studies of infection mechanisms, our aim was to immortalize bovine organospecific endothelial cells but also to assess their permissivity to viral infection. Using transfection with SV40 large T antigen, six bovine microvascular endothelial cell lines from various organs and one macrovascular cell line from an umbilical cord were established. They display their own panel of endothelial progenitor/mature markers, as assessed by flow cytometry and RT-qPCR, as well as the typical angiogenesis capacity. Using both Bluetongue and foot-and-mouth disease viruses, we demonstrate that some cell lines are preferentially infected. In addition, they can be transfected and are able to express viral proteins such as BTV8-NS3. Such microvascular endothelial cell lines bring innovative tools for in vitro studies of infection by viruses or bacteria, allowing for the study of host-pathogen interaction mechanisms with the actual in vivo target cells. They are also suitable for applications linked to microvascularization, such as anti-angiogenic and anti-tumor research, growing fields in veterinary medicine.

Pathogenic Escherichia coli in Dogs Reveals the Predominance of ST372 and the Human-Associated ST73 Extra-Intestinal Lineages.

Escherichia coli is a ubiquitous commensal and pathogen that has also been recognized as a multi-sectoral indicator of antimicrobial resistance (AMR). Given that latter focus, such as on resistances to extended-spectrum cephalosporins (ESC) and carbapenems, the reported population structure of E. coli is generally biased toward resistant isolates, with sequence type (ST)131 being widely reported in humans, and ST410 and ST648 being reported in animals. In this study, we characterized 618 non-duplicate E. coli isolates collected throughout France independently of their resistance phenotype. The B2 phylogroup was over-represented (79.6%) and positively associated with the presence of numerous virulence factors (VFs), including those defining the extra-intestinal pathogenic E. coli isolates (presence of ≥2 VFs: papA, sfaS, focG, afaD, iutA, and kpsMTII) and those more specifically related to uropathogenic E. coli (cnf1, hlyD). The major STs associated with clinical isolates from dogs were by far the dog-associated ST372 (20.7%) and ST73 (20.1%), a lineage that had commonly been considered until now as human-associated. Resistance to ESC was found in 33 isolates (5.3%), along with one carbapenemase-producing isolate, and was mostly restricted to non-B2 isolates. In conclusion, the presence of virulent E. coli lineages may be the issue, rather than the presence of ESC-resistant isolates, and the risk of transmission of such virulent isolates to humans needs to be further studied.

Experimental Ixodes ricinus-Sheep Cycle of Anaplasma phagocytophilum NV2Os Propagated in Tick Cell Cultures.

The causative agent of tick-borne fever and human granulocytic anaplasmosis, Anaplasma phagocytophilum, is transmitted by Ixodes ricinus, and is currently considered an emerging disease throughout Europe. In this study, we established a model of A. phagocytophilum sheep infection and I. ricinus transmission using the European Norway variant 2 ovine strain (NV2Os) propagated in both IDE8 and ISE6 tick cells. Two sheep were inoculated with IDE8 tick cells infected with NV2Os. Both sheep developed A. phagocytophilum infection as determined by qPCR and PCR, the presence of fever 4 days post inoculation (dpi), the observation of morulae in granulocytes at 6 dpi, and the detection of A. phagocytophilum antibodies at 14 dpi. A. phagocytophilum was detected by PCR in skin, lung, small intestine, liver, spleen, uterus, bone marrow, and mesenteric lymph node from necropsies performed at 14 and 15 dpi. One sheep was infested during the acute phase of infection with I. ricinus nymphs from a pathogen-free colony. After molting, A. phagocytophilum transstadial transmission in ticks was validated with qPCR positive bacterial detection in 80% of salivary glands and 90% of midguts from female adults. Infected sheep blood collected at 14 dpi was demonstrated to be able to infect ISE6 tick cells, thus enabling the infection of two additional naive sheep, which then went on to develop similar clinical signs to the sheep infected previously. One of the sheep remained persistently infected until 115 dpi when it was euthanized, and transmitted bacteria to 70 and 2.7% of nymphs engorged as larvae during the acute and persistent infection stages, respectively. We then demonstrated that these infected nymphs were able to transmit the bacteria to one of two other naive infested sheep. As expected, when I. ricinus females were engorged during the acute phase of infection, no A. phagocytophilum transovarial transmission was detected. The development of this new experimental model will facilitate future research on this tick-borne bacterium of increasing importance, and enable the evaluation of any new tick/transmission control strategies.

Bartonella Infection in Stray Dogs from Central and Southern Chile (Linares and Puerto Montt).

Bartonellae are emerging zoonotic vector-borne pathogens causing a broad spectrum of clinical symptoms in humans and animals, including life-threatening endocarditis. Dogs are infected with a wide range of Bartonella species and infection has been reported in free-roaming dogs from various South American countries. We report a high Bartonella seroprevalence in 82 Chilean stray dogs. More than half of the dogs from Linares (72.7%, n = 66) and Puerto Montt (56.2%, n = 16) were seropositive for Bartonella henselae, Bartonella vinsonii ssp. berkhoffii, or Bartonella clarridgeiae with antibody titers ranging from 1:64 to 1:512. Three dogs (3.6%) were PCR positive for Bartonella sp. Partial sequencing of the gltA gene indicated that two dogs were infected with B. henselae, and one with a strain close to Bartonella vinsonii ssp. vinsonii. Exposure to Bartonella species was common in stray Chilean dogs, as for other South American countries, likely associated with heavy ectoparasite infestation.

Genomic characterisation of a multidrug-resistant TEM-52b extended-spectrum ß-lactamase-positive Escherichia coli ST219 isolated from a cat in France.

OBJECTIVES: TEM-52 extended-spectrum ß-lactamases (ESBLs) have been detected in members of the Enterobacteriaceae isolated from human and non-human reservoirs, mainly in European countries. Here we report the first draft genome of a multidrug-resistant TEM-52b-positive Escherichia coli isolated from a companion animal in France. METHODS: Whole genomic DNA from E. coli 39590 was extracted and was sequenced using an Illumina NextSeq platform. De novo genome assembly was performed using Velvet v.1.2.10 and the draft genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline v.3.2. Genomic analyses were performed through bioinformatics tools from the Center for Genomic Epidemiology. RESULTS: The genome size was calculated as 5362108bp, with 5268 protein-coding sequences and a GC content of 50.5%. E. coli strain 39590 belonged to ST219, serotype O4:H34 and phylogroup E. The antimicrobial resistome consisted of genes encoding resistance to ß-lactams (blaTEM-52b), aminoglycosides [aph(3″)-Ib, aph(6)-Id, aadA2, aadA24], phenicols (catA1), sulfonamides (sul1, sul2), trimethoprim (dfrA1, dfrA14), lincosamides (lnuG) and tetracycline (tetA) as well as mutations in gyrA (Ser83Leu, Asp87Asn) and parC (Ser80Ile) conferring resistance to quinolones. Virulome analysis revealed iss, astA and eilA genes, and IncQ1, IncX4, IncX1, IncFIB and IncFIC plasmid incompatibility groups were identified. CONCLUSION: This draft genome can be used as a reference sequence for comparative studies using human and non-human E. coli isolates to identify genetic events that have contributed to pathogenicity and adaptation of TEM-52-producing E. coli clones at the human-animal interface as well as to elucidate dynamics of the spread of blaTEM-52 ESBL genes.

Supposed endogenous endophthalmitis caused by Serratia marcescens in a cat.

An 8-year-old male neutered domestic shorthair cat was presented for evaluation of acute respiratory distress. Respiratory auscultation revealed a diffuse and symmetric increase in bronchovesicular sounds. Thoracic radiographs showed a diffuse unstructured interstitial pulmonary pattern with multifocal alveolar foci. Despite an aggressive treatment with supportive care, including oxygenotherapy and systemic antibiotics, progressive respiratory distress increased. Three days after the presentation, acute anterior uveitis was noticed on left eye. Ophthalmic examination and ocular ultrasonography revealed unilateral panuveitis with ocular hypertension. The right eye examination was unremarkable. Cytological examination of aqueous humor revealed a suppurative inflammation. Serratia marcescens was identified from aqueous humor culture. Primary pulmonary infection was suspected but was not confirmed as owners declined bronchoalveolar lavage. Active uveitis resolved and cat's pulmonary status improved after appropriate systemic antibacterial therapy. Vision loss was permanent due to secondary mature cataract. To the best of authors' knowledge, this is the first report of endogenous bacterial endophthalmitis secondary to S. marcescens infection in a cat.

Frequency of bacteriuria in dogs with chronic kidney disease: A retrospective study of 201 cases.

BACKGROUND: Studies have shown an increased prevalence of positive urine culture (PUC) in cats with chronic kidney disease (CKD); no information is available in dogs. OBJECTIVES: To document the PUC frequency in a cohort of dogs with CKD, determine risk factors for PUC, and identify associations between clinicopathologic data and PUC. ANIMALS: Two hundred one client-owned dogs with CKD. METHODS: Retrospective, observational study. Dogs recruited from 2 veterinary teaching hospitals were included if they were diagnosed with CKD and had a culture performed on urine collected by cystocentesis. The PUC frequency was calculated, multivariate analysis was performed to identify risk factors, and associations with clinicopathologic data were investigated. RESULTS: Sixty-five dogs (32%) with CKD had PUC, including 8 (28%) in International Renal Interest Society (IRIS) stage 1; only 8% showed signs of a urinary tract infection. Escherichia coli was the most common isolate (67%). A PUC was more likely in females (odds ratio [OR], 3.22; 95% confidence interval [CI], 1.67-6.37; P < .001) than males and in dogs with isosthenuria (OR, 2.48; 95% CI, 1.24-5.03; P = .01) than in dogs with urine-specific gravity 1.013-1.024. A positive leukocyte esterase test and microorganisms found by urine sediment analysis were significantly associated with PUC (both P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Dogs with CKD, even IRIS stage 1, have a high frequency of PUC and most cases are asymptomatic. A urine culture could be considered in the routine evaluation of dogs with CKD, but the clinical relevance of a PUC remains unknown and needs further evaluation.

Management of Polyarthritis Affecting Sea Turtles at Kélonia, the Reunion Island Sea Turtle Observatory (2013-17).

All sea turtle species are listed on the Red List of the International Union for the Conservation of Nature due to multiple threats. Among these, disease is a frequent cause of stranded sea turtles encountered in rehabilitation centers. Since 2013, we found joint swelling in 13 sea turtles belonging to all four sea turtle species submitted to the Kélonia Sea Turtle Observatory of Reunion Island, France. Affected sea turtles presented with lameness, anorexia, and lethargy. Polyarthritis was radiographically confirmed and lesions were characterized by progressive osteolysis of bones surrounding joints. Anterior flippers were affected in all cases and posterior flippers were also involved in some cases. We isolated several bacterial agents from blood and synovial fluid. We attempted a collective treatment with injectable florfenicol, based on sensitivity results, which was continued for 4 wk and then as needed based on radiographic evolution of the lesions. Radiographic stabilization of the lesions occurred in nine of 13 cases. We reviewed environmental conditions and optimized them to minimize stress that could predispose these rehabilitated sea turtles to opportunistic infections. Handling techniques used to move sea turtles were also improved. While we can make no conclusion regarding the cause of polyarthritis in this population of sea turtles, we successfully managed this poorly described problem in a rehabilitation setting.

Bilateral bullous keratopathy secondary to melting keratitis in a Suri alpaca (Vicugna pacos).

An young alpaca was evaluated for bilateral progressive melting corneal ulcers and developped secondary bullous keratopathy during hospitalization. The tragic progression of melting ulcers in both eyes observed in our case leads us to recommend a rapid intensive medical therapy in young and debilitated alpacas presenting a corneal ulcer.

Co-circulation of different A. phagocytophilum variants within cattle herds and possible reservoir role for cattle.

BACKGROUND: Anaplasma phagocytophilum is a zoonotic tick-borne intracellular alpha-proteobacterium causing tick-borne fever, which leads to significant economic losses in domestic ruminants in Europe. Its epidemiological cycles are complex and reservoir host species of bovine strains have not yet been identified. Given that little genetic information is available on strains circulating within a defined bovine environment, our objective was to assess the genetic diversity of A. phagocytophilum obtained from the same farms over time. METHODS: Blood samplings were performed several times in two European herds. In the French herd, 169 EDTA-blood samples were obtained from 115 cows (32 were sampled two to four times). In the German herd, 20 cows were sampled six times (120 EDTA-blood samples). The presence of A. phagocytophilum DNA was assessed using a qPCR targeting msp2. The positive DNA samples underwent MLST at nine genetic markers (typA, ctrA, msp4, pleD, recG, polA, groEL, gyrA, and ankA). For each locus, sequences were aligned with available bacterial sequences derived from cattle, horse, dog, and roe deer hosts, and concatenated neighbor joining trees were constructed using three to six loci. RESULTS: Around 20% (57/289) of samples were positive. Forty positive samples from 23 French and six German cows (11 of them being positive at two time points) were sequenced. Six loci (typA, ctrA, msp4, pleD, recG, and polA) allowed to build concatenated phylogenetic trees, which led to two distinct groups of bovine variants in the French herd (hereafter called A and B), whereas only group A was detected in the German herd. In 42% of French samples, double chromatogram peaks were encountered in up to four loci. Eleven cows were found infected three weeks to 17 months after first sampling and harboured a new variant belonging to one or the other group. CONCLUSIONS: Our results demonstrate the occurrence of two major bovine strain groups and the simultaneous infection of single cows by more than one A. phagocytophilum strain. This challenges the role of cattle as reservoirs for A. phagocytophilum. This role may be facilitated via long-term bacterial persistence in individual cows and active circulation at the herd scale.