Fluorescent tagging for environmental surface cleaning surveillance in a veterinary hospital.

OBJECTIVES: To evaluate the use of fluorescent tagging for environmental surface cleaning surveillance in a small animal veterinary hospital and identify factors associated with tag removal. MATERIALS AND METHODS: Over 5.5 weeks, a commercial fluorescent dye (Glo Germ) was used to tag (mark) surfaces in a small animal veterinary teaching hospital. Twenty-four hours after tagging, cleaning was assessed with a black light (UV-A source). Surfaces were recorded as cleaned based on complete removal of fluorescent tagging at assessment. Proportions cleaned were calculated overall and by predictors (i.e. surface location/type, primary nature of surface contact - animal/human, week of study). RESULTS: A total of 4984 surfaces were tagged and assessed. Overall cleaning was 50%. Cleaning varied by surface/object (range: 2 to 100%) and hospital location (4 to 78%). Surfaces designated as having primarily animal contact were cleaned more frequently than those with primarily human contact (75%, 42%; P<0.001). Cleaning varied over the study period (range by week: 45 to 54%;); a significant trend was not identified. CLINICAL SIGNIFICANCE: Key surfaces in the small animal veterinary practice environment are unlikely to be adequately cleaned, posing a concern for animal and human health. Commercial products can be effectively used to asses environmental cleaning with findings used to target clinic-specific barriers to improve cleaning and reduce hospital-associated infections.

Molecular epidemiological analysis to assess the influence of pet-ownership in the biodiversity of Staphylococcus aureus and MRSA in dog- and non-dog-owning healthy households.

It has been suggested that zoonotic transmission of Staphylococcus aureus (SA) and methicillin-resistant S. aureus (MRSA) can occur between owners and their pets within the same household. However, the influence that pet-ownership could have in the biodiversity of SA/MRSA strains circulating among owners is not fully understood. The objective of this study was to perform a molecular epidemiological analysis to evaluate and compare the biodiversity of SA/MRSA strains in dog-owning and non-dog-owning healthy households within the same community. Antimicrobial resistance, SCCmec type, USA type and clonality were assessed. Overall, 33·1% (165/499) of human subjects carried SA and 2·8% (14/499) carried MRSA. Among dogs, 7·1% (8/113) carried SA but none were MRSA positive. No difference was detected in the diversity index of SA/MRSA pulsotypes between dog-owning and non-dog-owning households; but, a marked variation was still observed in the pulsotypes circulating in each type of household. Additionally, simultaneous carriage of the same SA pulsotype in owner(s) and dog was observed in 57% of households with positive humans and pets. These results demonstrate that dogs can indeed participate in the circulation of SA/MRSA pulsotypes within a home and that the presence of a pet does not seem to favour certain strains within their household.

Genetic relatedness and virulence factors of bovine Staphylococcus aureus isolated from teat skin and milk.

The objective of this study was to assess the role of teat skin colonization in Staphylococcus aureus intramammary infections (IMI) by evaluating genetic relatedness of Staph. aureus isolates from milk and teat skin of dairy cows using pulsed-field gel electrophoresis and characterizing the isolates based on the carriage of virulence genes. Cows in 4 known Staph. aureus-positive herds were sampled and Staph. aureus was detected in 43 quarters of 20 cows, with 10 quarters positive in both milk and skin (20 isolates), 18 positive only in milk, and 15 only on teat skin. Quarters with teat skin colonized with Staph. aureus were 4.5 times more likely to be diagnosed with Staph. aureus IMI than quarters not colonized on teat skin. Three main clusters were identified by pulsed-field gel electrophoresis using a cutoff of 80% similarity. All 3 clusters included both milk and skin isolates. The majority of isolates (72%) belonged to one predominant cluster (B), with 60% of isolates in the cluster originating from milk and 40% from teat skin. Genotypic variability was observed within 10 pairs (formed by isolates originating from milk and teat skin of the same quarter), where isolates in 5 out of the 10 pairs belonged to the same cluster. Forty-two virulence factors were screened using PCR. Some virulence factors were carried more frequently by teat skin isolates than by milk isolates or isolates from quarters with high somatic cell counts. Isolates in the predominant cluster B carried virulence factors clfA and clfB significantly more often than isolates in the minor clusters, which may have assisted them in becoming predominant in the herds. The present findings suggest that teat skin colonization with Staph. aureus can be an important factor involved in Staph. aureus IMI.

Comparative performance of contact plates, electrostatic wipes, swabs and a novel sampling device for the detection of Staphylococcus aureus on environmental surfaces.

AIMS: To evaluate the performance of four sampling methods [contact plates, electrostatic wipes (wipe), swabs and a novel roller sampler] for recovery of Staphylococcus aureus from a stainless steel surface. METHODS AND RESULTS: Stainless steel test plates were inoculated with Staph. aureus, dried for 24 h and sampled using each of the four methods. Samples were either incubated directly (roller, contact plate) or processed using elution and membrane filtration (swab, wipe). Performance was assessed by calculating the apparent sampling efficiency (ASE), analytical sensitivity (Sn) and percentage of replications with positive growth. The wipe demonstrated the best performance across all inoculating concentrations (ASE(48 h) = 18%; Sn(48 h) = 7 CFU per 100 cm(2)). The swab performed well when corrected for area actually sampled (ASE(48 h) = 24%; Sn(48 h) = 76 CFU per 100 cm(2)). Of the contact-based methods, the newly developed roller sampler outperformed the contact plate (roller: ASE(48 h) = 10%; Sn(48 h) = 17 CFU per 100 cm(2); contact plate: ASE(48 h) = 0·04%; Sn(48 h) = 1412 CFU per 100 cm(2)); both contact samplers performed better at higher inoculating concentrations (6E3 CFU per 100 cm(2) for the roller and 6E6 CFU per 100 cm(2) for the contact plate). Overall, the electrostatic wipe produced the highest number of replications resulting in positive growth (74%(24 h), 91%(48 h)). CONCLUSIONS: This study demonstrates that selection of the sampling method must be carefully considered, given that different methods have varying performance. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study assessing static wipes for sampling and one that uses a more real-world-relevant 24-h drying time. The results help with infection control, and environmental health professionals choose better sampling methodologies.

The use of plastic cover sheaths at the time of artificial insemination improved fertility of lactating dairy cows.

An adequate and clean artificial insemination (AI) technique is recommended to improve reproductive outcomes in dairy cattle. The objective of this study was to evaluate the effectiveness of using protective plastic sheaths (PS) to minimize contamination of the AI catheter (AIC) on pregnancies per AI (PAI) in lactating dairy cattle. Lactating cows housed in freestall barns on a commercial dairy farm were presynchronized with 2 injections of PGF(2α) given 14 d apart (starting at 26±3 d postpartum) followed by Ovsynch (GnRH-7 d-PGF(2α)-56 h-GnRH-16 h-timed-AI; TAI) 12 d later. Cows presenting signs of standing heat any time during the protocol received AI, whereas the remaining animals were subjected to TAI 16h after second Ovsynch GnRH. At the time of AI (1 AI technician), 996 services from 773 lactating dairy cows were randomly assigned to 1 of the 2 groups; with (TRT, n=487) or without (CON, n=509) the use of disposable PS. In the TRT group, the AIC protected with a PS was introduced into the vagina; once in the cranial portion of the vagina adjacent to the cervical os, the PS was pulled back and only the AIC was manipulated through the cervix into the uterine body for semen deposition. In the CON group, cows were inseminated without the use of PS. Samples were taken with a sterile cotton swab from the tip of the AIC (n=51) after AI from both treatment groups. Pregnancy diagnosis was determined by ultrasonography 39±3 d after AI. Cultured swab samples revealed that the use of PS was effective in minimizing contamination of the AIC (positive bacterial growth: TRT=61.53% vs. CON=100%). Overall, the proportion of cows pregnant was greater for cows in TRT (42.7) compared with the CON group (36.1). For first services postpartum, PAI did not differ between CON (43.01%, n=194) and TRT (43.8%, n=182) groups. However, PAI for second or greater services were greater in TRT (43.8%, n=305) than in CON cows (32.3%, n=315). Results from this study provided evidence that the use of PS during AI improved PAI for second or greater services in lactating dairy cows. Performing a clean AI technique through the use of PS may be an effective strategy to improve reproductive outcomes in dairy cattle.

Serological evidence of Neospora caninum in dual-purpose cattle herds in Venezuela.

Bovine neosporosis is a parasitic disease produced by Neospora caninum that induces abortion in cows, and consequently has a negative impact on the herd's reproductive efficiency. The main objective of this research was to determine the serological evidence of N. caninum in cattle herds from Venezuela using an indirect antibody capture ELISA test. Four hundred and fifty-nine (459) serum samples from crossbred adult cows were collected to be tested for Neospora antibodies. The sampled cows came from 15 large farms located in eight important cattle states that have predominant dual-purpose production systems (cattle from these farms are used for both milk and meat production). Fifty-two cows (11.3%) were seropositive to N. Caninum. Thirteen (86.7%) of 15 studied herds had cows seropositive to N. caninum. The average within-herd seroprevalence was 11.5% (range 3.8-36.7%). Cows that aborted in some of these farms had 2.71 (P: 0.009) greater odds to be seropositive when compared to cows that did not abort. Each one of the eight states represented in our study had seropositive animals. These results are the first evidence of exposure to N. caninum in Venezuelan cattle herds, indicating the possible circulation of this pathogen in the country. Further epidemiological studies should be granted to determine the spread of the disease in the Venezuelan cattle industry and its associated risk factors.

Reverse transcription-PCR assays for detection of bovine enteric caliciviruses (BEC) and analysis of the genetic relationships among BEC and human caliciviruses.

Two genetically distinct bovine enteric caliciviruses (BECs) have been identified: the norovirus (NLV) Jena and Newbury Agent-2 (NA-2) BECs, which are genetically related to human noroviruses, and the Nebraska (NB) BECs, which is related to sapoviruses and lagoviruses but may also represent a new calicivirus genus. The prevalence of these two BEC genotypes in cattle is unknown. Although reverse transcription-PCR (RT-PCR) primers for human NLV recognize NLV-BECs, the genetic relationships between NLV from humans and the NLV-BECs commonly circulating in cattle is undefined. In the present study, veal calf fecal samples were assayed for enteric caliciviruses by using six RT-PCR primer sets designed for the detection of human NLVs or BECs. Caliciviruses genetically related to the NLV-BEC Jena and NA-2 strains or to the recently characterized NB BEC strain were identified in three of four and four of four sampled veal herds, respectively. Extended 3'-terminal genome sequences of two NLV-BECs, designated CV95-OH and CV186-OH, encoding the RNA-dependent RNA polymerase (RdRp; open reading frame 1 [ORF-1]), VP1 (ORF-2), and VP2 (ORF-3) genes were determined. Phylogenetic and sequence identity analyses of each genome region demonstrated these viruses to be most closely related to the NLV-BEC Jena and NA-2 strains. In initial testing, the human P289-P290 (P289/290) primer set was found to be the most sensitive for calicivirus detection. However, its failure to identify all positive fecal pools (as determined by other assays) led us to design two new primer sets, CBECU-F/R and NBU-F/R, for the sensitive and specific detection of NLV-BEC (NLV-BEC Jena and NA-2) and BEC-NB-like viruses, respectively. The RT-PCR assays with the new primers were compared against other primer sets, including P289/290. Composite results of the tests completed by using the new assays identified 72% (54 of 75) of veal calf fecal samples as positive, with 21 of 21 sequenced reaction products specific for the target RdRp gene. The same design strategy used for the new BEC assays may also be applicable to the design of similar assays for the detection of human caliciviruses (HuCVs). Our data support the genetic relationship between NLV-BECs and NLV-HuCVs but with the NLV-BECs comprising two clusters within a third NLV genogroup.

Evaluation of concurrent shedding of bovine coronavirus via the respiratory tract and enteric route in feedlot cattle.

OBJECTIVE: To assess the relationship between shedding of bovine coronavirus (BCV) via the respiratory tract and enteric routes and the association with weight gain in feedlot cattle. ANIMALS: 56 crossbred steers. PROCEDURES: Paired fecal samples and nasal swab specimens were obtained and were tested for BCV, using antigen-capture ELISA. Paired serum samples obtained were tested for antibodies to BCV, using antibody-detection ELISA. Information was collected on weight gain, clinical signs, and treatments for enteric and respiratory tract disease during the study period. RESULTS: Number of samples positive for bovine respiratory coronavirus (BRCV) or bovine enteric coro navirus (BECV) was 37/224 (17%) and 48/223 (22%), respectively. Some cattle (25/46, 45%) shed BECV and BRCV. There were 25/29 (86%) cattle positive for BECV that shed BRCV, but only 1/27 (4%) cattle negative to BECV shed BRCV. Twenty-seven of 48 (56%) paired nasal swab specimens and fecal samples positive for BECV were positive for BRCV. In contrast, only 10/175 (6%) paired nasal swab specimens and fecal samples negative for BECV were positive for BRCV. Only shedding of BECV was associated with significantly reduced weight gain. Seroconversion to BCV during the 21 days after arrival was detected in 95% of the cattle tested. CONCLUSIONS AND CLINICAL IMPLICATIONS: Feedlot cattle infected with BCV after transport shed BCV from the respiratory tract and in the feces. Fecal shedding of BCV was associated with significantly reduced weight gain. Developing appropriate control measures for BCV infections could help reduce the decreased weight gain observed among infected feedlot cattle.