Development of an electronic interface for transfer of antimicrobial administration data in dairy farms.

Surveillance of antimicrobial administration in livestock production is an important factor in global policies to reduce spreading of antimicrobial resistance. In recent years, many studies have been carried out concerning the usage of antimicrobials in animal production and in some countries recording of antimicrobial quantities dispensed to famers is mandatory. On cattle farms, antimicrobial treatments are recorded for fattening calves under 8 months of age and for fattening cattle older than 8 months in Germany and treatment frequencies are then calculated. However, with the entry into force of Regulation (EU) 2019/6 on 01/28/2022, antimicrobial monitoring will gradually be extended to all animal species and age groups. Therefore, an effective, fast and accurate transfer of data on the use of antimicrobials, especially in the field of livestock farming, into corresponding databases is required to determine the treatment frequencies for the individual animal species or types of use. For this purpose, an electronic interface was programmed to transfer the data on antimicrobial use in dairy cattle farms from a herd management software program directly into a database. To test the practicability and effectiveness of this interface, 10 dairy cattle farms from Saxony were initially selected. Based on an in-depth analysis of the treatment frequencies of antimicrobial administration of 7 different age groups of animals after a two-year observation period, the functionality of the electronic interface could be established. The greatest potential for reduction of antimicrobials is in newborn calves, as they represent the age group with the highest treatment frequency.

Prevalence of Neutralizing Antibodies to Canine Distemper Virus and Response to Vaccination in Client-Owned Adult Healthy Dogs.

Re-vaccinations against canine distemper virus (CDV) are commonly performed in 3-year intervals. The study's aims were to determine anti-CDV antibodies in healthy adult dogs within 28 days of vaccination against CDV, and to evaluate factors associated with the presence of pre-vaccination antibodies and with the antibody response to vaccination. Ninety-seven dogs, not vaccinated within 1 year before enrollment, were vaccinated with a modified live CDV vaccine. A measurement of the antibodies was performed before vaccination (day 0), on day 7, and 28 after the vaccination by virus neutralization. A response to vaccination was defined as a ≥4-fold titer increase by day 28. Fisher's exact test was used to determine factors associated with a lack of antibodies and vaccination response. In total, 94.8% of the dogs (92/97; CI 95%: 88.2-98.1) had antibodies (≥10) prior to vaccination. A response to vaccination was not observed in any dog. Five dogs were considered humoral non-responders; these dogs neither had detectable antibodies before, nor developed antibodies after vaccination. Young age (<2 years) was significantly associated with a lack of pre-vaccination antibodies (p = 0.018; OR: 26.825; 95% CI: 1.216-1763.417). In conclusion, necessity of re-vaccination in adult healthy dogs should be debated and regular vaccinations should be replaced by antibody detection.

Multidrug-resistant enterobacteria in newborn dairy calves in Germany.

We studied the prevalence of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae in dairy calves as part of a routine health check protocol. In addition, data regarding antimicrobial use (AMU), farm hygiene, and farm management were collected in order to identify possible risks for ESBL occurrence. Ten farms participated in the study with a median of 781 milking cows (319-1701). All calves investigated were younger than two weeks with an average age of 6.8 (±3.9) days. The farms were visited and samples were collected twice at an interval of 7-11 months. Faecal samples diluted 1:10, were plated onto BrillianceTM ESBL agar in duplicates. After 24 hours at 37°C, colonies were counted and total colony forming units (cfu)/ml calculated. Bacteria species were identified biochemically. ESBL-production was phenotypically confirmed using the MICRONAUT-S ß-Lactamases system. Additionally, antimicrobial susceptibility was tested using VITEK® 2 technology. Phylotyping of E. coli isolates and screening for bla genes was performed by PCR. ESBL-producing enterobacteria were detected on all farms and 96.5% of calves investigated shed ESBL-positive bacteria. Of all ESBL-producing isolates, the majority were E. coli (92.9%), followed by Enterobacter cloacae (5.1%) and Klebsiella pneumoniae subsp. pneumoniae (2.0%). The majority of E. coli isolates was clearly assigned to phylogroup C (25.0%), followed by phylogroups A (15.2%) and E (14.1%). CTX-M group 1 was most frequently detected (80.4%). E. cloacae contained blaCTX-M and blaTEM or blaSHV. K. pneumoniae harboured blaSHV only. Besides resistance to penicillins and cephalosporins, the majority of isolates was also resistant to one or more antibiotic classes, with a high proportion being resistant against fluoroqinolones. 52.5% of isolates were further characterised as threefold multidrug resistant gram-negative bacteria (3MDR-GNB) according to the German Commission for Hospital Hygiene and Infection Prevention. None of the isolates were 4MDR-GNB, i.e. none revealed carbapenem-resistance. Penicillins were the most frequently administered antibiotics to calves on most farms and were the predominant substance class at herd level on all farms. Overall, the number of calves treated prior to sampling was rather low (11.7%). Analyses of data regarding the farm management identified weaknesses in biosecurity and cleaning and disinfection. Besides beta-lactam antibiotics being the most commonly used antibiotics no other risk factors could be identified. In summary, the prevalence of ESBL-carriers in dairy calves was exceptionally high and should be motivation to develop strategies for the reduction of multidrug-resistant bacteria in farm animals.

Antibody Response to Canine Parvovirus Vaccination in Dogs with Hypothyroidism Treated with Levothyroxine.

(1) Background: No information is available on how dogs with hypothyroidism (HypoT) respond to vaccination. This study measured pre- and post-vaccination anti-canine parvovirus (CPV) antibodies in dogs with HypoT treated with levothyroxine and compared the results to those of healthy dogs. (2) Methods: Six dogs with HypoT and healthy age-matched control dogs (n = 23) were vaccinated against CPV with a modified-live vaccine. Hemagglutination inhibition was used to measure antibodies on days 0, 7, and 28. The comparison of the vaccination response of dogs with HypoT and healthy dogs were performed with univariate analysis. (3) Results: Pre-vaccination antibodies (≥10) were detected in 100% of dogs with HypoT (6/6; 95% CI: 55.7-100) and in 100% of healthy dogs (23/23; 95% CI: 83.1-100.0). A ≥4-fold titer increase was observed in none of the dogs with HypoT and in 4.3% of the healthy dogs (1/23; CI95%: <0.01-22.7). Mild vaccine-associated adverse events (VAAEs) were detected in 33.3% of the dogs with HypoT (2/6; 95% CI: 9.3-70.4) and in 43.5% (10/23; 95% CI: 25.6-63.2) of the healthy dogs. (4) Conclusions: There was neither a significant difference in the dogs' pre-vaccination antibodies (p = 1.000), or vaccination response (p = 0.735), nor in the occurrence of post-vaccination VAAEs (p = 0.798). The vaccination response in dogs with levothyroxine-treated HypoT seems to be similar to that of healthy dogs.

Evaluation of a Point-of-Care Test for Pre-Vaccination Testing to Detect Antibodies against Canine Adenoviruses in Dogs.

(1) Background: Antibody testing is commonly used to assess a dog's immune status. For detection of antibodies against canine adenoviruses (CAVs), one point-of-care (POC) test is available. This study assessed the POC test´s performance. (2) Methods: Sera of 198 privately owned dogs and 40 specific pathogen-free (SPF) dogs were included. The reference standard for detection of anti-CAV antibodies was virus neutralization (VN) using CAV-1 and CAV-2 antigens. Specificity, sensitivity, positive predictive value (PPV), negative predictive value (NPV), and overall accuracy (OA) of the POC test were assessed. Specificity was considered most important. (3) Results: Prevalence of CAV-1 neutralizing antibodies (≥10) was 76% (182/238) in all dogs, 92% (182/198) in the subgroup of privately owned dogs, and 0% (0/40) in SPF dogs. Prevalence of CAV-2 neutralizing antibodies (≥10) was 76% (181/238) in all dogs, 91% (181/198) in privately owned dogs, and 0% (0/40) in SPF dogs. Specificity for detection of CAV-1 antibodies was lower (overall dogs, 88%; privately owned dogs, 56%; SPF dogs, 100%) compared with specificity for detection of CAV-2 antibodies (overall dogs, 90%; privately owned dogs, 65%; SPF dogs, 100%). (4) Conclusions: Since false positive results will lead to potentially unprotected dogs not being vaccinated, specificity should be improved to reliably detect anti-CAV antibodies that prevent infectious canine hepatitis in dogs.

Influence of Salmonella specific bacteriophages (O1; S16) on the shedding of naturally occurring Salmonella and an orally applied Salmonella Eastbourne strain in bearded dragons (Pogona vitticeps).

This study determined the passage time and phage propagation time of salmonella specific phages, Felix O1 and S16, in 10 bearded dragons, based on re-isolation from cloacal swabs and faecal samples following oral administration, as a possible tool for reducing salmonella shedding. In Study 1, Felix O1 was administered orally for 12 consecutive days. Over 60 days, swabs were taken from the oral cavity and cloaca and qualitative Salmonella detection as well as salmonella quantification from faecal samples were performed. In Study 2, a phage cocktail (Felix O1 and S16) was administered to half of the tested animals. Salmonella (S.) Eastbourne was also given orally to all animals. Oral and cloacal swabs were tested as in Study 1, and faecal samples were collected for phage quantification. Various Salmonella serovars were detectable at the beginning of the study. The numbers of serovars detected declined over the course of the study. S. Kisarawe was most commonly detected. Salmonella titres ranged from 102 to 107  cfu/g faeces. The phages (Felix O1 and S16) were detectable for up to 20 days after the last administration. The initial phage titres ranged from 103 to 107  pfu/ml. The study shows that the phages were able to replicate in the intestine, and were shed for a prolonged period and therefore could contribute to a reduction of Salmonella shedding.

Comparison of Four Commercially Available Point-of-Care Tests to Detect Antibodies against Canine Parvovirus in Dogs.

Measuring antibodies to evaluate dogs' immunity against canine parvovirus (CPV) is useful to avoid unnecessary re-vaccinations. The study aimed to evaluate the quality and practicability of four point-of-care (POC) tests for detection of anti-CPV antibodies. The sera of 198 client-owned and 43 specific pathogen-free (SPF) dogs were included; virus neutralization was the reference method. Specificity, sensitivity, positive and negative predictive value (PPV and NPV), and overall accuracy (OA) were calculated. Specificity was considered to be the most important indicator for POC test performance. Differences between specificity and sensitivity of POC tests in the sera of all dogs were determined by McNemar, agreement by Cohen's kappa. Prevalence of anti-CPV antibodies in all dogs was 80% (192/241); in the subgroup of client-owned dogs, it was 97% (192/198); and in the subgroup of SPF dogs, it was 0% (0/43). FASTest® and CanTiCheck® were easiest to perform. Specificity was highest in the CanTiCheck® (overall dogs, 98%; client-owned dogs, 83%; SPF dogs, 100%) and the TiterCHEK® (overall dogs, 96%; client-owned dogs, 67%; SPF dogs, 100%); no significant differences in specificity were observed between the ImmunoComb®, the TiterCHEK®, and the CanTiCheck®. Sensitivity was highest in the FASTest® (overall dogs, 95%; client-owned dogs, 95%) and the CanTiCheck® (overall dogs, 80%; client-owned dogs, 80%); sensitivity of the FASTest® was significantly higher compared to the one of the other three tests (McNemars p-value in each comparison: <0.001). CanTiCheck® would be the POC test of choice when considering specificity and practicability. However, differences in the number of false positive results between CanTiCheck®, TiterCHEK®, and ImmunoComb® were minimal.

Surgical hand preparation in an equine hospital: Comparison of general practice with a standardised protocol and characterisation of the methicillin-resistant Staphylococcus aureus recovered.

Presurgical hand asepsis is part of the daily routine in veterinary medicine. Nevertheless, basic knowledge seems to be low, even among specialised veterinary surgeons. The major objectives of our study were to assess current habits for presurgical hand preparation (phase 1) among personnel in a veterinary hospital and their effectiveness in reducing bacteria from hands in comparison to a standardised protocol (phase 2). Assessment of individual habits focused on time for hand washing and disinfection, the amount of disinfectant used, and the usage of brushes. The standardised protocol defined hand washing for 1 min with liquid neutral soap without brushing and disinfection for 3 min. All participants (2 surgeons, 8 clinic members, 32 students) used Sterillium®. Total bacterial counts were determined before and after hand washing, after disinfection, and after surgery. Hands were immersed in 100 ml sterile sampling fluid for 1 min and samples were inoculated onto Columbia sheep blood agar using the spread-plate method. Bacterial colonies were manually counted. Glove perforation test was carried out at the end of the surgical procedure. Differences in the reduction of relative bacterial numbers between current habits and the standardised protocol were investigated using Mann-Whitney-Test. The relative increase in bacterial numbers as a function of operation time (≤60 min, >60 min) and glove perforation as well as the interaction of both was investigated by using ANOVA. Forty-six and 41 preparations were carried out during phase 1 and phase 2, respectively. Individual habits differed distinctly with regard to time (up to 8 min) and amount of disinfectant (up to 48 ml) used both between participants and between various applications of a respective participant. Comparison of current habits and the standardised protocol revealed that the duration of hand washing had no significant effect on reducing bacteria. Contrary, the reduction in bacterial numbers after disinfection by the standardised protocol was significantly higher (p<0.001) compared to routine every-day practice. With regard to disinfection efficacy, the standardised protocol completely eliminated individual effects. The mean reduction in phase 1 was 90.72% (LR = 3.23; right hand) and 89.97% (LR = 3.28; left hand) compared to 98.85% (LR = 3.29; right hand) and 98.92% (LR = 3.47; left hand) in phase 2. Eight participants (19%) carried MRSA (spa type t011, CC398) which is well established as a nosocomial pathogen in veterinary clinics. The isolates could further be assigned to a subpopulation which is particularly associated with equine clinics (mainly t011, ST398, gentamicin-resistant). Glove perforation occurred in 54% (surgeons) and 17% (assistants) of gloves, respectively, with a higher number in long-term invasive procedures. Overall, bacterial numbers on hands mainly increased over time, especially when glove perforation occurred. This was most distinct for glove perforations on the left hand and with longer operating times. Our results demonstrate that standardised protocols highly improve the efficacy of hand asepsis measures. Hence, guiding standardised protocols should be prerequisite to ensure state-of-the-art techniques which is essential for a successful infection control intervention.

Diagnostic validation of a rapid and field-applicable PCR-lateral flow test system for point-of-care detection of cyprinid herpesvirus 3 (CyHV-3).

Koi herpesvirus disease (KHVD) is a highly infectious disease leading to outbreaks and mass mortality in captive and free-ranging common carp and koi carp. Outbreaks may result in high morbidity and mortality which can have a severe economic impact along the supply chain. Currently, control and prevention of KHVD relies on avoiding exposure to the virus based on efficient hygiene and biosecurity measures. An early diagnosis of the disease is crucial to prevent its spread and to minimize economic losses. Therefore, an easy-to-handle, sensitive, specific and reliable test prototype for a point-of-care detection of KHV was developed and evaluated in this study. We used a multiplex-endpoint-PCR followed by a specific probe hybridization step. PCR-products/hybridization-products were visualized with a simple and universal lateral flow immunoassay (PCR-LFA). Fifty-four gill tissue samples (KHV-positive n = 33, KHV-negative n = 21) and 46 kidney samples (KHV-positive n = 24, KHV-negative n = 22) were used to determine diagnostic sensitivity and specificity of the PCR-LFA. In addition, the usability of PCR-LFA to detect CyHV-3-DNA in gill swabs taken from 20 perished common carp during a KHVD-outbreak in a commercial carp stock was examined. This assay gave test results within approximately 60 min. It revealed a detection limit of 9 KHV gene copies/µl (95% probability), a diagnostic specificity of 100%, and diagnostic sensitivity of 94.81% if samples were tested in a single test run only. PCR inhibition was noticed when examining gill swab samples without preceding extraction of DNA or sample dilution. Test sensitivity coud be enhanced by examining samples in five replicates. Overall, our PCR-LFA proved to be a specific, easy-to-use and time-saving point-of-care-compatible test for the detection of KHV-DNA. Regarding gill swab samples, further test series using a higher number of clinical samples should be analyzed to confirm the number of replicates and the sample processing necessary to reveal a 100% diagnostic sensitivity.

Antibody Response to Canine Adenovirus-2 Virus Vaccination in Healthy Adult Dogs.

BACKGROUND: Re-vaccination against canine adenovirus (CAV) is performed in ≤3-year-intervals but their necessity is unknown. The study determined anti-CAV antibodies within 28 days of re-vaccination and factors associated with the absence of antibodies and vaccination response. METHODS: Ninety-seven healthy adult dogs (last vaccination ≥12 months) were re-vaccinated with a modified live CAV-2 vaccine. Anti-CAV antibodies were measured before vaccination (day 0), and after re-vaccination (day 7, 28) by virus neutralization. A ≥4-fold titer increase was defined as vaccination response. Fisher's exact test and multivariate regression analysis were performed to determine factors associated with the absence of antibodies and vaccination response. RESULTS: Totally, 87% of dogs (90/97; 95% CI: 85.61-96.70) had anti-CAV antibodies (≥10) before re-vaccination. Vaccination response was observed in 6% of dogs (6/97; 95% CI: 2.60-13.11). Time since last vaccination (>3-5 years, OR = 9.375, p = 0.020; >5 years, OR= 25.000, p = 0.006) was associated with a lack of antibodies. Dogs from urban areas were more likely to respond to vaccination (p = 0.037). CONCLUSION: Many dogs had anti-CAV pre-vaccination antibodies, even those with an incomplete vaccination series. Most dogs did not respond to re-vaccination. Based on this study, dogs should be re-vaccinated every 3 years or antibodies should be determined.

Antibody Response to Canine Parvovirus Vaccination in Dogs with Hyperadrenocorticism Treated with Trilostane.

It is unknown how dogs with hyperadrenocorticism (HAC) respond to vaccination. This study measured antibodies against canine parvovirus (CPV) in dogs with HAC treated with trilostane before and after CPV vaccination, and compared the immune response to that from healthy dogs. Eleven dogs with HAC, and healthy age-matched control dogs (n = 31) received a modified-live CPV vaccine. Antibodies were determined on days 0, 7, and 28 by hemagglutination inhibition. Univariate analysis was used to compare the immune response of dogs with HAC and healthy dogs. Pre-vaccination antibodies (≥10) were detected in 100% of dogs with HAC (11/11; 95% CI: 70.0-100) and in 93.5% of healthy dogs (29/31; 95% CI: 78.3-99.2). No ≥4-fold increase in antibody titer was observed in dogs with HAC while in 22.6% of healthy dogs, a ≥4-fold titer increase was observed (7/31; 95% CI: 11.1-40.1). Mild vaccine-associated adverse events (VAAEs) were detected in 54.5% of dogs with HAC (6/11; 95% CI: 28.0-78.8) and in 29.0% of healthy dogs (9/31; 95% CI: 15.9-46.8). There was neither a significant difference in presence of pre-vaccination antibodies (p = 1.000), or response to vaccination (p = 0.161), nor in the occurrence of VAAEs (p = 0.158). Immune function of dogs with HAC treated with trilostane seems comparable to that of healthy dogs.

Borderline resistance to oxacillin in Staphylococcus aureus after treatment with sub-lethal sodium hypochlorite concentrations.

Surface disinfectants are regularly used in prophylactic and infection control measures. Concern has been raised whether residues of sub-inhibitory disinfectant concentrations may constitute a selective pressure and could contribute to the development of strains which are tolerant and/or resistant to biocides including antibiotics. The current study investigated whether Staphylococcus (S.) aureus ATCC® 29213™ and ATCC® 6538™ would change their growth characteristics and antimicrobial susceptibility profiles after prolonged treatment with sub-inhibitory concentrations of sodium hypochlorite (NaOCl). NaOCl is a fast-acting disinfectant with a broad-spectrum activity, inexpensive and widely used in healthcare and the food production industry. Minimum inhibitory concentration (MIC) for NaOCl was determined by broth macrodilution according to the guidelines for disinfectant efficacy testing provided by the German Veterinary Medical Society. Serial passages after 24 h and 72 h, respectively, in defined sub-inhibitory concentrations of NaOCl resulted in a number of phenotypic variants. Two of these variants, derived from S. aureus ATCC® 29213™, showed elevated MICs of oxacillin and were considered as in vitro-generated borderline oxacillin-resistant S. aureus (BORSA). Transmission electron microscopy revealed a significantly thickened cell wall in these isolates, a phenomenon that has also been described for Listeria monocytogenes after low-level exposure to NaOCl. Whole genome sequencing revealed an early stop codon in the gene coding for the GdpP protein and thereby abolishing the function of this gene. GdpP represents a phosphodiesterase that regulates gene expression, and loss of function of the GdpP protein has been described in association with borderline oxacillin resistance. Our findings suggest that a mutation in the GdpP protein gene and morphological changes of the cell wall were induced by repeated exposure to sub-lethal NaOCl concentrations, and most likely accounted for a BORSA phenotype in two variants derived from S. aureus ATCC® 29213™.

Antibody response to feline herpesvirus-1 vaccination in healthy adult cats.

OBJECTIVES: Vaccination against feline herpesvirus-1 (FHV-1) is recommended for all cats. However, it is unknown how adult healthy cats with different pre-vaccination antibodies respond to FHV-1 vaccination in the field. The aim of the study was to determine the prevalence of neutralising antibodies against FHV-1 in healthy adult cats and the response to FHV-1 vaccination within 28 days of vaccination. METHODS: One hundred and ten cats (⩾1 year of age) that had not received a vaccination within the past 12 months were vaccinated with a combined FHV-1 vaccine. Antibodies against FHV-1 were determined before vaccination (day 0), on day 7 and day 28 by serum neutralisation test. Uni- and multivariate statistical analyses were used to determine factors associated with the presence of pre-vaccination antibodies and response to vaccination. RESULTS: Pre-vaccination neutralising antibody titres (⩾10) were present in 40.9% of cats (45/110; 95% confidence interval [CI] 32.2-50.3); titres were generally low (range 10-640). Antibody response to vaccination (⩾four-fold titre increase) was observed in 8.3% (9/109; 95% CI 4.2-15.1). Cats ⩾2 years of age were more likely to have pre-vaccination neutralising antibodies than cats aged between 1 and 2 years (odds ratio [OR] 24.619; P = 0.005). Cats from breeders were more likely to have pre-vaccination neutralising antibodies than privately owned cats (OR 7.070; P = 0.007). Domestic shorthair cats were more likely to have an at least four-fold titre increase vs purebred cats (OR 11.22; P = 0.027). CONCLUSIONS AND RELEVANCE: Many cats have no detectable neutralising antibodies against FHV-1 despite previous vaccinations and fail to develop a ⩾four-fold titre increase after vaccination. This is likely because older cats and cats with a higher FHV-1 exposure risk are more likely to get infected with FHV-1 and thus to have FHV-1 neutralizing antibodies. Purebred cats more often fail to develop a ⩾four-fold titre increase after vaccination.

Impact of UVC-sustained recirculating air filtration on airborne bacteria and dust in a pig facility.

High amounts of aerial pollutants like dust and microorganisms can pose serious health hazards to animals and humans. The aim of the current study therefore was, to assess the efficiency of UVC irradiation combined to air filtration in reducing airborne microorganisms at laboratory scale. In a second part, a UVC-combined recirculating air filtration module (UVC module) was implemented in a small animal facility in order to assess its improvement of air quality with regard to airborne bacteria and dust. Tests at laboratory scale were performed using aerosols of Staphylococcus (S.) aureus, Actinobacillus pleuropneumoniae, porcine parvovirus (PPV) and porcine reproductive and respiratory syndrome virus. We varied relative humidity (RH) to evaluate its effect on UVC irradiation efficiency. In addition, viability of pathogens inside the filter material was determined over up to six months. UVC-combined air filtration resulted in a more than 99% reduction of viral and bacterial particles. RH had no influence on UVC efficiency. Viability in the filter matter varied depending on the pathogen used and RH with S. aureus and PPV being most resistant. In our small pig facility consisting of two separated barns, weekly air measurements were conducted over a period of 13 weeks (10 piglets) and 16 weeks (11 piglets), respectively. Airborne bacterial numbers were significantly lower in the barn equipped with the UVC module compared to the reference barn. On average a reduction to 37% of reference values could be achieved for bacteria, whereas the amount of total dust was reduced to a much lesser extent (i.e. to 78% of reference values). Measures taken in front of and behind the UVC module revealed a reduction of 99.4% for airborne bacteria and 95.0% for total dust. To conclude, recirculating air filtration combined to UVC provided efficient reduction of pathogens at laboratory and experimental scale. The implementation of such devices might improve the overall environmental quality in animal facilities.

The use of a salmonella bacteriophage in bearded dragons: application, passage time and reisolation.

OBJECTIVE: This study determined the passage time and phage propagation time of a salmonella specific phage, Felix O1, in bearded dragons, based on reisolation from cloacal swabs and faecal samples following oral administration, as a possible tool for reducing the zoonotic risk of salmonella from pet reptiles. An application scheme for this phage in bearded dragons was developed. MATERIAL AND METHODS: Ten healthy bearded dragons (Pogona vitticeps) were used in the study. The pH tolerance of the phage was tested and drugs were used to evaluate their influence on the gastric pH of the reptiles. After pH adjustment, the phage was administered orally for 12 consecutive days. Over 60 days, swabs were taken from the cloaca and examined for the presence of phages using culture and PCR. Furthermore, faecal samples were collected for phage quantification. RESULTS: Felix O1 displayed no activity at pH below 2.8. A calcium- and magnesium carbonate buffer induced an appropriate gastric pH increase for 30 minutes. Phages were reisolated for up to 24 days (mean shedding: 19 days) after last administration. Titres between 105 and 107 plaque forming units/g faeces were detected. The animals did not show any clinical signs related to phage application. CONCLUSION AND CLINICAL RELEVANCE: The study provides first results on oral administration, passage time, and reisolation of a phage in reptiles. It could be shown that the phage was able to replicate in the intestine, and was shed for a prolonged period and therefore could potentially contribute to a reduction of salmonella shedding.

Antibody Response to Feline Calicivirus Vaccination in Healthy Adult Cats.

This study evaluated the prevalence of feline calicivirus (FCV) antibodies and response to vaccination in healthy adult cats. Cats >1 year (n = 111) that had not been vaccinated within 12 months of enrollment in the study received a vaccine containing inactivated FCV antigen strains 431 and G1. Antibodies were determined on Days 0, 7, and 28 by virus neutralization (VN) using FCV isolate KS20, and by broad spectrum blocking FCV enzyme-linked immunosorbent assay (ELISA). Factors associated with the presence of antibodies and vaccine response were determined by uni- and multivariate analysis. Pre-vaccination antibodies were detected in 62.2% of cats (CI95%: 52.9-70.1) by VN and in 77.2% (CI95%: 67.5-84.6) by ELISA. A ≥4-fold titer increase after vaccination was observed in 13.6% (CI95%: 8.3-21.4) of cats with VN and 33.7% (CI95%: 24.5-44.5) with ELISA. Factors associated with the presence of pre-vaccination VN antibodies were age (≥2 years; OR: 7.091; p = 0.022) and lack of previous vaccination (OR: 3.472; p = 0.014). The presence of pre-vaccination ELISA antibodies was associated with time since last vaccination (OR: 5.672; p = 0.043). Outdoor cats were more likely to have a ≥4-fold ELISA titer increase (OR: 5.556; p = 0.005). Many cats had pre-vaccination FCV antibodies, and their presence depended on previous vaccinations and increases with age. A ≥4-fold titer increase was rarely observed and was influenced by the lifestyle of the cat.

Faecal shedding of parvovirus deoxyribonucleic acid following modified live feline panleucopenia virus vaccination in healthy cats.

Positive canine parvovirus (CPV) faecal test results have been reported in dogs after modified live virus (MLV) vaccination. Thus, the aim was to investigate feline panleucopenia virus (FPV) shedding in recently vaccinated, adult, clinically healthy cats and to assess related factors. Forty cats were vaccinated with an FPV MLV vaccine. Faeces of cats were tested for presence of parvovirus DNA on days 7, 14, 21 and 28 by quantitative real-time PCR; DNA-positive samples were subjected to partial VP2 gene sequencing. Virus isolation was performed whenever sufficient amounts of faeces were available. Serum antibody titres were measured by haemagglutination inhibition on days 0, 7 and 28. Overall, 30.0 per cent (12/40; 95% CI 18.0 to 45.6) of cats shed parvovirus DNA. Sequencing revealed FPV vaccine virus DNA in three cats, FPV field virus DNA in four cats and CPV field virus DNA in one cat. Shedding was significantly associated with lack of prevaccination antibody titres (40) (P=0.016; OR: 6.44; 95% CI 1.44 to 28.89) and with postvaccination titre increases (fourfold) (P=0.029; OR: 5.00; 95% CI 1.17 to 21.39). Shedding of field or vaccine virus DNA seems to be common in healthy cats which can be a concern in shelters and catteries. Diagnostic tools should be developed to facilitate differentiation of vaccine and field virus shedding.

Evaluation of disinfectant efficacy against multidrug-resistant bacteria: A comprehensive analysis of different methods.

BACKGROUND: Multidrug-resistant gram-negative bacteria (MDR-GNB) constitute a threat to health care worldwide. Disinfectants are used to prevent and control the spread of MDR-GNB in a hospital setting but their efficacy might be impaired by bacterial mechanisms that may act on both antimicrobials and disinfectants. Determination of minimum inhibitory concentrations is mainly used to determine bacterial susceptibility against disinfectants, but practical tests on surfaces might be more suitable to predict in-use conditions. Our objective was to compare and evaluate 4 different methods widely used to assess surface disinfectant efficacy. METHODS: The efficacy of benzalkonium chloride (BAC), peracetic acid (PAA), and ethanol (ETH) against multidrug-resistant Acinetobacter, Pseudomonas, and Klebsiella strains was assessed by minimum inhibitory concentration determinations, quantitative suspension tests, qualitative suspension tests, and carrier tests. Test results were compared to ascertain the most appropriate method. RESULTS: ETH, PAA, and BAC were highly effective against MDR-GNB, but we observed marked differences in efficacious concentrations (up to 100-fold) as a function of the test method applied. Minimum inhibitory concentration determination was not reliable for evaluating susceptibility or resistance to BAC. CONCLUSIONS: Surface tests should be used to determine bacterial susceptibility against disinfectants. Moreover, suitable guidelines are needed that allow for the standardization and comparison of bactericidal values obtained by different investigators.

Antibody response to feline panleukopenia virus vaccination in cats with asymptomatic retrovirus infections: a pilot study.

OBJECTIVES: Currently, there are only a few studies on how immunocompromised cats, such as cats infected with feline leukaemia virus (FeLV) and feline immunodeficiency virus (FIV), respond to vaccination. Therefore, this study measured feline panleukopenia virus (FPV) antibodies in retrovirus-infected cats within a period of 28 days after FPV vaccination, and compared the immune response to that of non-infected cats. METHODS: Eight asymptomatic retrovirus-infected cats (four FeLV, four FIV), and non-infected age-matched control cats (n = 67) were vaccinated with a commercial FPV modified live virus (MLV). Pre- and post-vaccination antibody titres were measured by haemagglutination inhibition (HI) on days 0, 7 and 28. An HI titre ⩾1:40 was defined as protective. An adequate response to vaccination was defined as a four-fold titre increase or higher. Comparison of the immune response of retrovirus-infected and non-infected cats was performed. RESULTS: Pre-vaccination FPV antibody titres ⩾1:40 were present in 100% (n = 8/8; 95% confidence interval [CI] 62.8-100) of retrovirus-infected and in 77.6% (n = 52/67; 95% CI 66.2-86.0) of non-infected cats. An adequate response to vaccination (titre increase ⩾four-fold) was seen in 1/8 retrovirus-infected cats (12.5%; 95% CI 0.1-49.2) compared with 22/67 non-infected cats (32.8%; 95% CI 22.8-44.8). In cats with high pre-vaccination titres (⩾1:160), a four-fold titre increase or higher was observed in 1/8 retrovirus infected cats (12.5%; 95% CI 0.1-49.2) compared with 4/42 non-infected cats (9.5%; 95% CI 3.2-22.6). None of the eight retrovirus-infected cats developed illness or vaccination side effects after vaccination with MLV against FPV within the 28 days. There were no significant differences between groups: for pre-vaccination titres; for at least four-fold titre increases following vaccination in either all cats or the cats with high pre-vaccination titres; and concerning adverse effects. CONCLUSIONS AND RELEVANCE: All retrovirus-infected asymptomatic cats had pre-vaccination FPV antibodies indicating protection against panleukopenia. Response of retrovirus-infected cats to vaccination was similar to the response of non-infected cats.

Development and evaluation of a broth macrodilution method to determine the biocide susceptibility of bacteria.

In comparison to biocide efficacy testing, biocide susceptibility testing of bacteria so far lacks standardized methods for routine use. The aims of the present study were to develop a broth macrodilution method to test bacterial pathogens for their biocide susceptibility and to evaluate this method in an interlaboratory trial. Staphylococcus aureus ATCC®6538 was tested for its susceptibility to benzalkonium chloride, chlorhexidine and isopropanol comparing test strain suspension preparations, test volumes and incubation times. The use of 2 mL volumes for the testing and an incubation time of 24 h were proposed. Ten German laboratories participated in the interlaboratory trial. Four reference strains (S. aureus ATCC®6538, Enterococcus hirae ATCC®10541, Escherichia coli ATCC®10536 and Pseudomonas aeruginosa ATCC®15442) commonly used for biocide activity testing, were included. Strains were tested three times at independent occasions for their susceptibility to benzalkonium chloride, glutardialdehyde and isopropanol. In total, 360 data points were obtained (30 per strain/biocide combination). The modal minimal inhibitory concentration ± one dilution step was defined as acceptable range. For the four reference strains and the three biocides 80-100% of the values were considered as acceptable. The deviations within the laboratories for a strain/biocide combination were rather consistent. In general, the testing was performed without difficulties by the laboratories. Although inoculum plate counts of four laboratories were outside the acceptable range, this did not have a large impact on the results. The proposed method was stable and easy to perform. It may contribute to a harmonization and standardization of biocide susceptibility testing.