Variations in association of nasal microbiota with virulent and non-virulent strains of Glaesserella (Haemophilus) parasuis in weaning piglets

International audienceAbstractGlaesserella (formerly Haemophilus) parasuis causes Glässer’s disease, which results in high economic loss in the swine industry. To understand the polymicrobial interactions of G. parasuis and the nasal microbiota, the statistical association patterns of nasal colonizing bacteria with virulent and non-virulent strains of G. parasuis were studied accounting for the farm management practices as potential risk factors for the occurrence of Glässer’s disease. The nasal microbiota from 51 weaned-piglets from four farms with Glässer’s disease and three farms with no respiratory diseases was previously characterized and included in this study. The presence of virulent and/or non-virulent G. parasuis strains in the nasal cavities was determined in order to establish the potential association with other members of the nasal microbiota. Multivariate logistic and linear regression models were performed among the various members of nasal microbiota and G. parasuis. The multi-site production system and disease presence in the farm were both significantly associated with the presence of G. parasuis virulent strains in the nose of the piglets. Differential bacterial associations were observed with virulent or non-virulent G. parasuis. Chitinophagaceae, Corynebacteriaceae and Corynebacterium were positively associated with the virulent G. parasuis strains, while Enterobacteriaceae, Peptostreptococcaceae, Clostridium XI, and Escherichia/Shigella were negatively associated with virulent G. parasuis. On the other hand, Flavobacteriaceae, Planobacterium, and Phascolarctobacterium were positively associated with the non-virulent G. parasuis strains, while Rikenellaceae, Enterococcaceae, Odoribacter, and Corynebacterium were negatively associated with non-virulent G. parasuis. In conclusion, the nasal microbiota communities showed variations in the association with the G. parasuis strains type

Molecular detection of Treponema species organisms in foremilk and udder cleft skin of dairy cows with digital dermatitis

Identification of reservoirs and transmission routes of digital dermatitis (DD)-associated Treponema spp. is considered an effective means for controlling DD infection in dairy cows. The objective of this study is to identify and characterize the potential reservoir niches for DD-associated Treponema spp. from healthy udder cleft skin and foremilk in lactating dairy cows. A large dairy farm was visited weekly from March to July 2015. Clinical investigation revealed that a total of 25 lame cows had DD lesions located at the plantar aspect of the interdigital cleft. A total of 75 samples, three per cow, were collected including deep swabs from DD lesions (n = 25), non-aseptically collected foremilk samples (n = 25) and skin swabs from udder cleft (n = 25). Treponema spp. were identified using nested PCR assays and confirmed by DNA sequencing. Results revealed that Treponema phagedenis (T. phagedenis)-like was the most identified species in the foremilk 40% (10/25), in comparison with DD lesions and udder cleft skin samples with 32% (8/25) and 20% (5/25), respectively. On the other hand, Treponema pedis (T. pedis) was the most identified species in the udder cleft skin 80% (20/25), in comparison with DD lesions and foremilk samples with 68% (17/25) and 60% (15/25), respectively. None of the examined samples were identified by PCR as containing DNA from Treponema medium (T. medium) or Treponema vincentii (T. vincentii)-like. To the best of our knowledge, this is the first report for detection of T. phagedenis-like and T. pedis from healthy skin of udder cleft and foremilk samples. Detection of DD Treponema spp. from udder cleft skin and foremilk samples indicates that these sites could be potential reservoirs for spirochetes involved in DD. Udder cleft skin and foremilk may have a role in transmission routes of DD Treponema in dairy farms.info:eu-repo/semantics/acceptedVersio

Diagnostic performance of direct and indirect methods for assessing failure of transfer of passive immunity in dairy calves using latent class analysis

Accurate diagnosis of failure of transfer of passive immunity (FTPI) in newborn calves is an essential component of dairy farm management plan. Several methods (direct and indirect) are available for diagnosis of FTPI in dairy calves. However, the indirect methods offer an advantage over the direct methods in not requiring an experienced veterinarian, rapid, cost efficient and can be performed under field-setting. The objective of this study was to estimate the diagnostic performance of radial immunodiffusion (RID) assay, transmission infrared (TIR) spectroscopy and digital Brix refractometer for diagnosis of FTPI in dairy calves using latent class models at four cut-off values of digital Brix refractometer. Holstein calves (n = 691) from 40 commercial dairy farms in the four Atlantic Canada provinces were blood-sampled and tested for detection of FTPI. Results showed that the number of calves with FTPI was 253 (36.6%) by RID, 194 (28.1%) by TIR and 204 (29.5%) by Brix refractometer at cut-off value of 8.2%. Estimates of SeRID was higher than SeTIR and SeBrix, at all Brix refractometer cut-offs, but with increase of Brix refractometer cut-off from 8.2 to 8.5%, SeRID and SeTIR were decreased from 96.0% (95% PCI: 88.0–99.0) and 79.0% (95% PCI: 70.0–85.0), to 92.0% (95% PCI: 77.0–99.0) and 74.0% (95% PCI: 61.0–82.0), respectively. SpRID and SpTIR were always higher than SpBrix at all tested cut-offs and were above 92.0%, and 96.0%, respectively. With increasing the cut-off of Brix refractometer from 8.2 to 8.5%, SeBrix estimate has remarkably increased from 79.0% (95% PCI: 70.0–96.0) to 95.0% (95% PCI: 87.0–100.0), respectively. Whilst, SpBrix was decreased from 95.0% (95% PCI: 91.0–98.0) at cut-off 8.2% to 84.0% (95% PCI: 78.0–94.0) at cut-off 8.5%. In conclusion, RID has a higher Se than TIR and Brix, if the latter is used with cut-offs of 8.2% or 8.3%. However, the higher the cut-off, the more comparable sensitivities of RID and digital Brix refractometer. The median estimate of SpTIR was always higher than SpRID and SpBrix at all tested cut-offs. However, the 95% confidence interval estimates of the three tests were overlapping across the tested cut-offs of digital Brix refractometer reflecting the inability to prefer a test over the other based on the Sp estimate.info:eu-repo/semantics/acceptedVersio

Prevalence and Diffusion of Gastrointestinal Parasite Infections in Swamp Water Buffalo (Bubalus Bubalis) Populations from Marshlands of Iraq

Background and objectives: New information on the epidemiology of gastrointestinal (GI) parasite infection in water buffaloes is crucial for understanding their risk factors and transmission. The objectives of this study were (1) to determine the prevalence of GI parasites in buffaloes in the Marshland areas of southern Iraq, and (2) to evaluate the association of risk factors with the parasitic infections.Materials and Methods: A total of 166 water buffaloes from the Marshland in the north of Basra (n=75), and Thi-Qar (n=91) provinces from November 2016 to April 2017 were enrolled. Fecal samples were collected and examined for the presence of helminth eggs and protozoal oocysts using sedimentation-flotation and centrifugal flotation techniques.Results: The overall prevalence of infection in buffaloes was 82% (136/166), with the highest number of single parasite infection (64%), followed by those with double (29%) and triple (7%) parasite infections. The most frequently identified parasites were Fasciola spp. (23%, 39/166), Eimeria spp. (19%, 32/166), Toxocara vitulorum (13%, 21/166), Trichostrongylus spp. (12%, 20/166), and Oesophagostomum spp. (10%, 10/166). Moniezia spp. was the only identified cestode with a prevalence of (8%, 13/166). A significant association was reported between feeding type and parasitic infections with Eimeria spp., Trichostrongylus spp., Moniezia spp., Trichuris spp., and Ostertagia ostertagia.Conclusion: The prevalence of GI parasitic infection in buffaloes raised in the Marshlands is high, indicating a high intensity of natural infection. The findings of this study imply an urgent need for the implementation of efficient control measures against parasitic infections in the Marshlands

Communications of Staphylococcus aureus and non-aureus Staphylococcus species from bovine intramammary infections and teat apex colonization

The role of non-aureus staphylococci (NAS) in the risk of acquisition of intramammary infections with Staphylococcus aureus is vague and still under debate. The objectives of this study were to (1) investigate the distribution patterns of NAS species from milk and teat skin in dairy herds with automatic milking systems, and (2) examine if the isolated NAS influences the expression of S. aureus virulence factors controlled by the accessory gene regulator (agr) quorum sensing system. In 8 herds, 14 to 20 cows with elevated somatic cell count were randomly selected for teat skin swabbing and aseptic quarter foremilk samples from right hind and left front quarters. Teat skin swabs were collected using the modified wet-dry method and milk samples were taken aseptically for bacterial culture. Colonies from quarters with suspicion of having NAS in milk or teat skin samples (or both) were subjected to MALDI-TOF assay for species identification. To investigate the interaction between S. aureus and NAS, 81 isolates NAS were subjected to a qualitative β-galactosidase reporter plate assay. In total, 373 NAS isolates were identified representing 105 from milk and 268 from teat skin of 284 quarters (= 142 cows). Sixteen different NAS species were identified, 15 species from teat skin and 10 species from milk. The most prevalent NAS species identified from milk were Staphylococcus epidermidis (50%), Staphylococcus haemolyticus (15%), and Staphylococcus chromogenes (11%), accounting for 76%. Meanwhile, the most prevalent NAS species from teat skin were Staphylococcus equorum (43%), S. haemolyticus (16%), and Staphylococcus cohnii (14%), accounting for 73%. Using reporter gene fusions monitoring transcriptional activity of key virulence factors and regulators, we found that out of 81 supernatants of NAS isolates, 77% reduced expression of hla, encoding a-hemolysin, 70% reduced expression of RNAIII, the key effector molecule of agr, and 61% reduced expression of spa encoding protein A of S. aureus, respectively. Our NAS isolates showed 3 main patterns: (1) downregulation effect such as S. chromogenes (milk) and Staphylococcus xylosus (milk and teat), (2) no effect such as Staphylococcus sciuri (teat) and S. vitulinus (teat), and the third pattern (c) variable effect such as S. epidermidis (milk and teat) and S. equorum (milk and teat). The pattern of cross-talk between NAS species and S. aureus virulence genes varied according to the involved NAS species, habitat type, and herd factors. The knowledge of how NAS influences S. aureus virulence factor expression could explain the varying protective effect of NAS on S. aureus intramammary infections.info:eu-repo/semantics/acceptedVersio

Typeability of MALDI-TOF assay for identification of non-aureus staphylococci associated with bovine intramammary infections and teat apex colonization

Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF), a culture-dependent assay, has recently been implemented for routine identification of non-aureus staphylococci (NAS) species from milk, but the assay has never been investigated for NAS from nonmilk or environmental samples. The objective of this study was to evaluate the typeability of the MALDI-TOF assay for the identification and differentiation of bovine-associated NAS species on aseptically collected quarter milk and teat skin samples in dairy herds. In 8 herds, 14 to 20 cows with elevated somatic cell count were randomly selected for teat skin swabs and foremilk samples from right hind and left front quarters. Teat skin swabs and milk samples were collected aseptically for preliminary identification using bacterial culture on chromogenic and calf blood agars. Colonies from milk and teat skin samples with suspicion of having NAS were identified to species-level by MALDI-TOF assay. Out of 511 isolates from 284 quarters (142 cows), 78% (n = 399) were identified by MALDI-TOF. The percentage of correctly identified NAS from milk (91%, 105/115) using MALDI-TOF was higher than the percentage from teat skin (68%, 268/396). Out of the identified isolates, 93% (n = 373) were successfully identified as NAS, whereas the remaining 26 (7%) were shown to be other bacterial species. Out of 26 NAS isolates, 1 originated from milk (Corynebacterium stationis), whereas 25 originated from teat skin representing Aerococcus viridans (n = 7), Bacillus pumilus (n = 13), Enterococcus saccharolyticus (n = 1), Clostridium septicum (n = 1), Corynebacterium stationis (n = 2), and Corynebacterium casei (n = 1). The MALDI-TOF identified 85 (98/115) and 62% (245/396) of the isolates in the first test. Isolates that were not identified to species-level at first test were subjected to a second test, and 47 (8/17) and 32% (48/151) from milk and teat skin, respectively, were identified. After 2 rounds of MALDI-TOF, 22% (n = 112) of the isolates were not identified, representing 103 from teat skin and 9 from milk. Eighteen isolates without identification by MALDI-TOF were successfully identified to species-level using sequencing, where 16 were correctly identified as NAS, whereas the other 2 were Corynebacterium stationis. In conclusion, MALDI-TOF is a reliable assay for identification and typeability of NAS species from aseptically collected quarter milk samples. The assay may be used for identification of NAS species from teat skin swabs. However, confirmation using nucleic acid-based tools is vital for accurate species identification of some species and strains.info:eu-repo/semantics/acceptedVersio

Epidemiological and ultrasonographic investigation of bovine fascioliasis in smallholder production system in Eastern Nile Delta of Egypt

Regular updating of our knowledge on the epidemiological determinants of bovine fascioliasis is necessary to increase the awareness of the disease's significance and subsequently, improve the control measures. The objectives of this study were (1) to estimate the prevalence of bovine fascioliasis, and identify the association of epidemiological characteristics under traditional householders' production systems, (2) to describe the association between the clinical picture, Fasciola spp. egg count and hepatobiliary ultrasonography findings. In total, 270 faecal samples were examined microscopically for the presence or absence of Fasciola spp. egg, using the sedimentation-flotation method. Copro-positive animals were subjected to ultrasonographic examination. Overall prevalence of copro-positive animals was 27.4% (22.4-33.0%, 95% CI). The final multivariate analysis showed that there was a significant association between fascioliasis and animal species (P < 0.03), and administration of anthelmintic (P < 0.0001). Cattle have a less chance of being positive to Fasciola spp. by 0.55 (95% CI: 0.30 - 0.99) compared to water buffaloes. Administration of anthelmintic to animals on a regular basis decreased the risk of copro-positivity to Fasciola spp by 0.17 (95% CI: 0.07 - 0.36) compared to animals received anthelmintic on an irregular basis. Infected animals having different Fasciola spp. egg burden revealed different clinical symptoms associated with hepatobiliary changes on ultrasonographic examination ranged from normal hepatic parenchyma and bile system in low faecal egg load to hyperechogenic hepatic parenchyma, hyperechogenic with distal shadowing bile duct, and distended gallbladder in high faecal egg load of Fasciola spp. In conclusion, the prevalence of bovine fascioliasis is high under the traditional household's production system. Regular administration of anthelmintic significantly reduces the animal's chance of being copro-positive to Fasciola spp. Ultrasound poses a valuable prognostic technique for assessment of bovine fascioliasis.info:eu-repo/semantics/acceptedVersio

Immune responses following neonatal vaccination with conserved F4 fragment of VtaA proteins from virulent Glaesserella parasuis adjuvanted with CAF®01 or CDA

Glaesserella parasuis is a Gram-negative bacterium that colonizes the upper airways of swine, capable of causing a systemic infection called Glässer’s disease. This disease is more frequent in young post-weaning piglets. Current treatments against G. parasuis infection are based on the use of antimicrobials or inactivated vaccines, which promote limited cross-protection against different serovars. For this reason, there is an interest in developing novel subunit vaccines with the capacity to confer effective protection against different virulent strains. Herein, we characterize the immunogenicity and the potential benefits of neonatal immunization with two different vaccine formulations based on the F4 polypeptide, a conserved immunogenic protein fragment from the virulence-associated trimeric autotransporters of virulent G. parasuis strains. With this purpose, we immunized two groups of piglets with F4 combined with cationic adjuvant CAF®01 or cyclic dinucleotide CDA. Piglets immunized with a commercial bacterin and non-immunized animals served as control groups. The vaccinated piglets received two doses of vaccine, at 14 days old and 21 days later. The immune response induced against the F4 polypeptide varied depending on the adjuvant used. Piglets vaccinated with the F4+CDA vaccine developed specific anti-F4 IgGs, biased towards the induction of IgG1 responses, whereas no anti-F4 IgGs were de novo induced after immunization with the CAF®01 vaccine. Piglets immunized with both formulations displayed balanced memory T-cell responses, evidenced upon in vitro re-stimulation of peripheral blood mononuclear cells with F4. Interestingly, pigs immunized with F4+CAF®01 controlled more efficiently a natural nasal colonization by a virulent serovar 4 G. parasuis that spontaneously occurred during the experimental procedure. According to the results, the immunogenicity and the protection afforded by F4 depend on the adjuvant used. F4 may represent a candidate to consider for a Glässer‘s disease vaccine and could contribute to a better understanding of the mechanisms involved in protection against virulent G. parasuis colonization.This study was financially supported by the European Project TRANSVAC2–730964–INFRAIA–2016-1 of the European Vaccine Initiative funded in turn by the European Commission under the Horizon 2020 Program. Sergi López-Serrano was funded by this Project. IRTA-CReSA is also supported by the Centres de Recerca de Catalunya (CERCA) Program from the Generalitat de Catalunya.info:eu-repo/semantics/publishedVersio

Association between teat skin colonization and intramammary infection with Staphylococcus aureus and Streptococcus agalactiae in herds with automatic milking systems

The objective of this study was to investigate the association between teat skin colonization and intramammary infection (IMI) with Staphylococcus aureus or Streptococcus agalactiae at the quarter level in herds with automatic milking systems. Milk and teat skin samples from 1,142 quarters were collected from 300 cows with somatic cell count >200,000 cells/mL from 8 herds positive for Strep. agalactiae. All milk and teat skin samples were cultured on calf blood agar and selective media. A subset of samples from 287 quarters was further analyzed using a PCR assay (Mastit4 PCR; DNA Diagnostic A/S, Risskov, Denmark). Bacterial culture detected Staph. aureus in 93 (8.1%) of the milk samples and 75 (6.6%) of the teat skin samples. Of these, 15 (1.3%) quarters were positive in both the teat skin and milk samples. Streptococcus agalactiae was cultured in 84 (7.4%) of the milk samples and 4 (0.35%) of the teat skin samples. Of these, 3 (0.26%) quarters were positive in both the teat skin and milk samples. The PCR detected Staph. aureus in 29 (10%) of the milk samples and 45 (16%) of the teat skin samples. Of these, 2 (0.7%) quarters were positive in both the teat skin and milk samples. Streptococcus agalactiae was detected in 40 (14%) of the milk samples and 51 (18%) of the teat skin samples. Of these, 16 (5.6%) quarters were positive in both the teat skin and milk samples. Logistic regression was used to investigate the association between teat skin colonization and IMI at the quarter level. Based on bacterial culture results, teat skin colonization with Staph. aureus resulted in 7.8 (95% confidence interval: 2.9; 20.6) times higher odds of Staph. aureus IMI, whereas herd was observed as a major confounder. However, results from the PCR analyses did not support this association. Streptococcus agalactiae was isolated from the teat skin with both PCR and bacterial culture, but the number of positive teat skin samples detected by culture was too low to proceed with further analysis. Based on the PCR results, Strep. agalactiae on teat skin resulted in 3.8 (1.4; 10.1) times higher odds of Strep. agalactiae IMI. Our results suggest that Staph. aureus and Strep. agalactiae on teat skin may be a risk factor for IMI with the same pathogens. Focus on proper teat skin hygiene is therefore recommended also in AMS.info:eu-repo/semantics/publishedVersio