Flies (Insecta, Diptera) collected in the environment of dairy farms as carriers of Rotavirus A and betacoronavirus.

AIMS: We aimed to investigate the prevalence of rotavirus and coronavirus in dipterans that commonly inhabit the environment of dairy farms. METHODS AND RESULTS: We collected 217 insect specimens from nine dairy farms, which were examined through hemi-nested RT-PCR followed by Sanger sequencing in search of VP1 and N genes for rotavirus and bovine coronavirus-BCoV, respectively. With a predominance of Muscidae (152/217 = 70%) 11 families of Diptera were identified. Rotavirus A (RVA) and betacoronavirus (BCoV) were detected in 14.7% (32/217) and 4.6% (10/217) of the dipterans, respectively. Sequencing of the amplicons was possible for 11.5% (25/217) of RVA and 0.5% (1/217) of BCoV, confirming the presence of these pathogens. CONCLUSIONS: Our findings highlight the role of dipterans as carriers of RVA and BCoV of great relevance for public and animal health.

Genome-Based Characterization of Multidrug-Resistant Escherichia coli Isolated from Clinical Bovine Mastitis.

Mastitis occurrence in dairy cows is a broad topic that involves several sectors, from antimicrobial resistance and virulence of strains to economic implications and cattle management practices. Here, we assessed the molecular characterization (antimicrobial resistance determinants, virulence genes, sequences type, serotypes, and plasmid types) of 178 Escherichia coli strains isolated from milk samples from cows with clinical mastitis using a genome-based k-mers approach. Of these, 53 (29.8%) showed multidrug resistance by disc diffusion. We selected eight multidrug-resistant mastitis-associated E. coli for whole-genome sequencing and molecular characterization based on raw data using k-mers. We assessed antimicrobial resistance genes, virulence factors, serotypes, Multilocus Sequence Typing (MLST), and plasmid types. The most antimicrobial resistance gene found were blaTEM-1B (7/8), tetA (6/8), strA (6/8), strB (6/8), and qnrB19 (5/8). A total of 25 virulence factors were detected encoding adhesins, capsule, enzymes/proteins, increased serum survival, hemolysin, colicins, and iron uptake. These virulence factors were associated with Extraintestinal Pathogenic E. coli. Three pandemic clones were found: ST10, ST101, and ST69. Two E. coli were assigned in the O117 serogroup and one in the O8:H25 serotype. The most common plasmid groups were IncFII (7/8) and IncFIB (6/8). Our findings contribute to the knowledge of virulence mechanisms, epidemiological aspects, and antimicrobial resistance determinants of E. coli strains obtained from clinical mammary infections of cows.

Biofilm production under different atmospheres and growth media by Streptococcus agalactiae isolated from milk of cows with subclinical mastitis.

Different methods to analyze Streptococcus agalactiae biofilm formation have been investigated, but standardized protocols have not been developed. We compared S. agalactiae biofilm production among different atmospheres and growth media. Biofilm formation was studied in 32 isolates from bovine mastitis cases grown in Tryptone Soy Broth (TSB), Todd Hewitt Broth (THB), Luria Bertani Broth (LB) and Brain Heart Infusion (BHI), under two atmospheres, aerobic and 5% CO2. Regardless of the culture medium, growth under 5% CO2 resulted in a greater proportion of biofilm formation (65.63%), as compared with aerobic conditions (39.84%). Regardless of the atmosphere, the chances of biofilm formation were greater for isolates grown in TSB, as compared with THB [Odds ratio (OR) = 3.02], BHI (OR = 4.57), or LB (OR = 10.20). Thus, we suggest the use of 5% CO2 atmosphere and TSB in biofilm formation assays by Group-B streptococci (GBS) isolated from intramammary infections.

Refinement and partial validation of the UNESP-Botucatu multidimensional composite pain scale for assessing postoperative pain in horses.

BACKGROUND: Quantification of pain plays a vital role in the diagnosis and management of pain in animals. In order to refine and validate an acute pain scale for horses a prospective, randomized, blinded study was conducted. Twenty-four client owned adult horses were recruited and allocated to one of four following groups: anaesthesia only (GA); pre-emptive analgesia and anaesthesia (GAA,); anaesthesia, castration and postoperative analgesia (GC); or pre-emptive analgesia, anaesthesia and castration (GCA). One investigator, unaware of the treatment group, assessed all horses at time-points before and after intervention and completed the pain scale. Videos were also obtained at these time-points and were evaluated by a further four blinded evaluators who also completed the scale. The data were used to investigate the relevance, specificity, criterion validity and inter- and intra-observer reliability of each item on the pain scale, and to evaluate construct validity and responsiveness of the scale. RESULTS: Construct validity was demonstrated by the observed differences in scores between the groups, four hours after anaesthetic recovery and before administration of systemic analgesia in the GC group. Inter- and intra-observer reliability for the items was only satisfactory. Subsequently the pain scale was refined, based on results for relevance, specificity and total item correlation. CONCLUSIONS: Scale refinement and exclusion of items that did not meet predefined requirements generated a selection of relevant pain behaviours in horses. After further validation for reliability, these may be used to evaluate pain under clinical and experimental conditions.

Dermatological and morphological findings in quarter horses with hereditary equine regional dermal asthenia.

BACKGROUND: Hereditary equine regional dermal asthenia (HERDA) is an autosomal recessive disorder affecting quarter horses (QHs); affected horses exhibit characteristic skin abnormalities related to abnormal collagen biosynthesis. HYPOTHESIS/OBJECTIVES: To characterize the thickness and morphological abnormalities of the skin of HERDA-affected horses and to determine the interobserver agreement and the diagnostic accuracy of histopathological examination of skin biopsies from horses with HERDA. ANIMALS: Six affected QHs, confirmed by DNA testing, from a research herd and five unaffected QHs from a stud farm. METHODS: The skin thickness in 25 distinct body regions was measured on both sides in all affected and unaffected horses. Histopathological and ultrastructural evaluation of skin biopsies was performed. RESULTS: The average skin thickness in all of the evaluated regions was thinner in the affected horses. A statistically significant difference between skin thickness of the affected and unaffected animals was observed only when the average magnitude of difference was ≥38.7% (P = 0.038). The interobserver agreement for the histopathological evaluation was fair to substantial. The histopathological sensitivity for the diagnosis of HERDA was dependent on the evaluator and ranged from 73 to 88%, whereas the specificity was affected by the region sampled and ranged from 35 to 75%. CONCLUSIONS AND CLINICAL IMPORTANCE: Despite the regional pattern of the cutaneous signs, skin with decreased thickness was not regionally distributed in the HERDA-affected horses. Histopathological evaluation is informative but not conclusive for establishing the diagnosis. Samples of skin from the neck, croup or back are useful for diagnosis of HERDA. However, the final diagnosis must be confirmed using molecular testing.

Validation of a Questionnaire for Equine Personality Assessement.

We propose a practical method of assessing the personality of horses using five personality axes. Four evaluators empirically judged 19 horse individuals on specific adjectives for each axis. To validate the questionnaire, four behavioral tests were conducted with these same animals (social interactions, novel object test, bridge test,and arena test). In this tests, the frequency of specific behaviors were evaluated to create a scale related to the same personality adjectives and judge the animals based on their reactions.The questionnaire was reliable in determining the personality of horses, since the results were consistent with those obtained through behavioral tests. Additionally, in this group of horses attention reactions were more frequent than fear reactions, but significant differences occurred among tests. This study proposes a practical questionnaire for owners and trainers to assess the personality of their horses. The application of this tool can improve the relationship between humans and horses, directing a more empathic approach in the everyday routine with these animals.

Long-Term Co-Circulation of Host-Specialist and Host-Generalist Lineages of Group B Streptococcus in Brazilian Dairy Cattle with Heterogeneous Antimicrobial Resistance Profiles.

Group B Streptococcus (GBS) is a major cause of contagious bovine mastitis (CBM) in Brazil. The GBS population is composed of host-generalist and host-specialist lineages, which may differ in antimicrobial resistance (AMR) and zoonotic potential, and the surveillance of bovine GBS is crucial to developing effective CBM control and prevention measures. Here, we investigated bovine GBS isolates (n = 156) collected in Brazil between 1987 and 2021 using phenotypic testing and whole-genome sequencing to uncover the molecular epidemiology of bovine GBS. Clonal complex (CC) 61/67 was the predominant clade in the 20th century; however, it was replaced by CC91, with which it shares a most common recent ancestor, in the 21st century, despite the higher prevalence of AMR in CC61/67 than in CC91, and high selection pressure for AMR from indiscriminate antimicrobial use in the Brazilian dairy industry. CC103 also emerged as a dominant CC in the 21st century, and a considerable proportion of herds had two or more GBS strains, suggesting poor biosecurity and within-herd evolution due to the chronic nature of CBM problems. The majority of bovine GBS belonged to serotype Ia or III, which was strongly correlated with CCs. Ninety-three isolates were resistant to tetracycline (≥8 µg/mL; tetO = 57, tetM = 34 or both = 2) and forty-four were resistant to erythromycin (2.0 to >4 µg/mL; ermA = 1, ermB = 38, mechanism unidentified n = 5). Only three isolates were non-susceptible to penicillin (≥8.0 µg/mL), providing opportunities for improved antimicrobial stewardship through the use of narrow-spectrum antimicrobials for the treatment of dairy cattle. The common bovine GBS clades detected in this study have rarely been reported in humans, suggesting limited risk of interspecies transmission of GBS in Brazil. This study provides new data to support improvements to CBM and AMR control, bovine GBS vaccine design, and the management of public health risks posed by bovine GBS in Brazil.

Serological study of vaccinia virus reservoirs in areas with and without official reports of outbreaks in cattle and humans in São Paulo, Brazil.

Vaccinia virus (VACV), the etiological agent of an exanthematic disease, has been associated with several bovine outbreaks in Brazil since the end of the global vaccination campaign against smallpox. It was previously believed that the vaccine virus used for the WHO global campaign had adapted to an unknown wild reservoir and was sporadically re-emerging in outbreaks in cattle and milkers. At present, it is known that Brazilian VACV is phylogenetically different from the vaccinia virus vaccinal strain, but its origin remains unknown. This study assessed the seroprevalence of orthopoxviruses in domestic and wild animals and farmers from 47 farms in three cities in the southwest region of the state of São Paulo with or without official reports of outbreaks in cattle or humans. Our data indicate a low seroprevalence of antibodies in wild animals and raise interesting questions about the real potential of wild rodents and marsupials as VACV reservoirs, suggesting other routes through which VACV can be spread.

Occurrence of mycobacteria in bovine milk samples from both individual and collective bulk tanks at farms and informal markets in the southeast region of Sao Paulo, Brazil.

BACKGROUND: Mycobacterium spp. is one of the most important species of zoonotic pathogens that can be transmitted from cattle to humans. The presence of these opportunistic, pathogenic bacteria in bovine milk has emerged as a public-health concern, especially among individuals who consume raw milk and related dairy products. To address this concern, the Brazilian control and eradication program focusing on bovine tuberculosis, was established in 2001. However, bovine tuberculosis continues to afflict approximately 1,3 percent of the cattle in Brazil. In the present study, 300 samples of milk from bovine herds, obtained from both individual and collective bulk tanks and informal points of sale, were cultured on Löwenstein-Jensen and Stonebrink media. Polymerase chain reaction (PCR)-based tests and restriction-enzyme pattern analysis were then performed on the colonies exhibiting phenotypes suggestive of Mycobacterium spp., which were characterized as acid-fast bacilli. RESULTS: Of the 300 bovine milk samples that were processed, 24 were positively identified as Mycobacterium spp.Molecular identification detected 15 unique mycobacterial species: Mycobacterium bovis, M. gordonae, M. fortuitum, M. intracellulare, M. flavescens, M. duvalii, M. haemophilum, M. immunogenum, M. lentiflavum, M. mucogenicum, M. novocastrense, M. parafortuitum, M. smegmatis, M. terrae and M. vaccae. The isolation of bacteria from the various locations occurred in the following proportions: 9 percent of the individual bulk-tank samples, 7 percent of the collective bulk-tank samples and 8 percent of the informal-trade samples. No statistically significant difference was observed between the presence of Mycobacterium spp. in the three types of samples collected, the milk production profiles, the presence of veterinary assistance and the reported concerns about bovine tuberculosis prevention in the herds. CONCLUSION: The microbiological cultures associated with PCR-based identification tests are possible tools for the investigation of the presence of Mycobacterium spp. in milk samples. Using these methods, we found that the Brazilian population may be regularly exposed to mycobacteria by consuming raw bovine milk and related dairy products. These evidences reinforces the need to optimize quality programs of dairy products, to intensify the sanitary inspection of these products and the necessity of further studies on the presence of Mycobacterium spp. in milk and milk-based products.

Enterotoxin genes in coagulase-negative and coagulase-positive staphylococci isolated from bovine milk.

The objective of this study was to isolate and identify the main staphylococcal species causing bovine mastitis in 10 Brazilian dairy herds and study their capability to produce enterotoxins. Herds were selected based on size and use of milking technology, and farms were visited once during the study. All mammary glands of all lactating cows were screened using the California Mastitis Test (CMT) and a strip cup. A single aseptic milk sample (20 mL) was collected from all CMT-positive quarters. Identification of Staphylococcus spp. was performed using conventional microbiology, and PCR was used to determine the presence of enterotoxin-encoding genes (sea, seb, sec, and sed). Of the 1,318 CMT-positive milk samples, Staphylococcus spp. were isolated from 263 (19.9%). Of these isolates, 135 (51%) were coagulase-positive staphylococci (CPS) and 128 (49%) were coagulase-negative staphylococci (CNS). Eighteen different species of CNS were isolated, among which S. warneri, S. epidermidis and S. hyicus were the most frequent. The distribution of Staphylococcus species was different among herds: S. epidermidis was found in 8 herds, S. warneri was found in 7 herds, and S. hyicus in 6 herds. Some of the CNS species (S. saprophyticus ssp. saprophyticus, S. auricularis, S. capitis, and S. chromogenes) were isolated in only one of the farms. Genes related to production of enterotoxins were found in 66% (n=85) of all CNS and in 35% of the CPS isolates. For both CNS and CPS isolates, the most frequently identified enterotoxin genes were sea, seb, and sec; the prevalence of sea differed between CPS (9.5%) and CNS (35.1%) isolates. Staphylococcus warneri isolates showed a greater percentage of sea than seb, sec, or sed, whereas S. hyicus isolates showed a greater percentage of sea than sec. Over 60% of CNS belonged to 3 major species, which carried 62.2 to 81.3% of the enterotoxin genes. The high prevalence highlights the potential for food poisoning caused by these species. For possible high-risk situations for food poisoning, such as milk produced with total bacterial counts greater than regulatory levels and stored under inappropriate temperatures, monitoring contamination with CNS could be important to protect human health. Because the prevalence of CNS intramammary infections in dairy herds is usually high, and these species can be found in great numbers in bulk milk, identification of risk factors for production of staphylococcal enterotoxins should be considered in future studies.

Relationship between Biofilm Production and High Somatic Cell Count in Streptococcus agalactiae Isolated from Milk of Cows with Subclinical Mastitis.

Streptococcus agalactiae (S. agalactiae) is one of the main agents that causes mastitis in dairy cows, mainly inducing the subclinical form, which is characterized by a high somatic cell count (SCC). The aim of this study was to correlate the increase in SCC caused by S. agalactiae in cows with subclinical mastitis to the presence of genes related to adhesion and invasion in bovine mammary epithelial cells (BMEC) and biofilm formation. Considering the 145 isolates tested, 57.2% presented the capsular type Ia and 42.8% presented type III. We identified the virulence genes among the isolates and determined nine genetic profiles. The most common profile was identified in 69 isolates (47.5%): Ia, fbsA+, fbsB-, pI1-, pI2a-, pI2b+, and hylb+. All isolates produced biofilm, with 58.6% classified as strong producers, 29% as moderate producers and 12.4% as weak producers. No statistical correlation was found between the presence of virulence genes and increased SCC or biofilm production. However, biological evidence was observed between increased SCC and biofilm production. One isolate from each profile was randomly subjected to adhesion and invasion assays, and all of them adhered to BEMC, but none were able to invade. Our results showed that different genetic profiles do not provide advantages for bacteria to invade BMEC in vitro. In addition, biofilm production appears to be related to high SCC.

In Vitro Activity of Quaternary Ammonium in Prototheca Isolated from Clinical Bovine Mastitis Identified by Mass Spectrometry and PCR Sequencing of the cytb Gene Marker.

The in vitro algaecide activity of quaternary ammonium (QA) against Prototheca isolated from bovine clinical mastitis was investigated, in which the clinical severity was scored, milk samples were subjected to microbiological culture, and algal species were identified by molecular typing. A total of 4275 milk clinical samples of different cows from ten large dairy farms were used. Forty-four (1%) samples of cows from three dairy farms yielded growth of Prototheca, of which 88.6% (39/44) were identified as Prototheca bovis and 11.3% (5/44) as Prototheca sp. by MALDI-TOF MS, whereas 100% of the isolates were identified as P. bovis using PCR sequencing of the cytb gene. Among cows for which clinical severity scoring was available, 78.8% (26/33) and 21.2% (7/33) had mild and moderate infections, respectively, whereas no animal showed severe clinical signs. The algaecide activity of QA in Prototheca was observed in low concentrations among all isolates, in 20.4% (9/44) at 35 ppm, 36.4% (16/44) at 17 ppm, and 43.2% (19/44) at an 8 ppm, in addition to activity on three reference Prototheca strains. Overall, the study highlights the predominance of P. bovis as the causative agent of algal mastitis in bovines. Prototheca induced abnormalities preponderantly in the milk and mammary gland tissue of cows, and to our knowledge, our study is the first to apply clinical severity scoring in protothecal mastitis. In addition, the study underlines the activity of QA in low concentrations against Prototheca, indicating its potential use as an antiseptic/disinfectant in milking facilities and dairy environments.

Sequential Cerebrospinal Fluid Sampling in Horses: Comparison of Sampling Times and Two Different Collection Sites.

Analysis of the cerebrospinal fluid (CSF) is important as a complementary test in horses with neurologic diseases, and sequential analysis may provide information about the treatment response or evolution and quantitative measures of the CSF drug concentration during treatment. The aim of this study was to compare erythrocyte and nucleated cell counts and protein concentration in multiple CSF samples obtained sequentially from two different puncture sites in clinically healthy horses. Eight and 12 horses, with no evidence of neurologic disease, were subjected to CSF collection from the atlanto-occipital (AO) and C1-C2 spaces, respectively. Cytologic and chemical analyses were performed on the CSF obtained at five sampling times (T1, T2, T3, T4, and T5). Repeated measures models were used to compare the mean erythrocyte count, nucleated cell count, and total protein concentration between the AO and C1-C2 groups at each sampling time. C1-C2 CSF had a significantly higher total protein concentration at T1 and T4 than that of AO CSF. All total protein concentration values remained within the reference interval (<90 mg/dL) for all sampling times and groups. No statistical difference was present between results at T2, T3, T4, and T5 and at T1 in both groups for all analyses. In conclusion, five consecutive AO or C1-C2 CSF collections with at least a 7-d interval did not result in alterations in the CSF erythrocyte and nucleated cell counts and total protein concentrations and did not interfere with the CSF analysis results.

Epidemiological aspects of Ancylostoma spp. infection in naturally infected dogs from São Paulo state, Brazil.

Ancylostoma spp. are the most prevalent canine nematode parasites in Brazil. Despite their widespread parasitism in juvenile dogs, infections may occur regardless of host age. Although eosinophilia is a frequent finding in experimental infections, it is uncommon in naturally infected dogs. This study aimed to evaluate the prevalence of Ancylostoma spp. and the eosinophil blood counts (EBC) in naturally infected dogs, with or without comorbidities, admitted to the Veterinary Teaching Hospital of the São Paulo State University (UNESP), School of Veterinary Medicine and Animal Science, Botucatu campus, São Paulo state, Brazil, from 2009 to 2018. All retrospective data were gathered from veterinary medical records (VMR); diagnosis of Ancylostoma infection required the identification of eggs in fecal samples by the simple flotation test (SFT). Ancylostoma-infected animals were evaluated for other intestinal parasitic coinfections (IPC) by either the SFT or the centrifugal-flotation test. Dogs free of any gastro-intestinal parasites were prospectively included in control group (Group C). Ancylostoma-infected animals were defined: Ancylostoma spp. only intestinal parasite infection (Group A), Ancyslostoma spp. with concurrent IPC (Group B), Ancylostoma spp. only intestinal parasite infection with concurrent systemic disorders (Group D), and Ancylostoma spp. with both IPC and concurrent systemic disorders (Group E). The overall prevalence of Ancylostoma spp. was 12.1% (207/1715), that was decreased from 2014 to 2018 (9.7%) relative to the 2009-to-2013 period (13.9%). Prevalence was not significantly different between dogs <1-year-old (10.7% [51/478]) and ≥ 1 year-old (11.7% [130/1109]). IPC was observed in 45.4% (93/205) of the animals positive for Ancylostoma spp., while dogs <1 year old experienced IPC more often (58.8% [30/51]) than dogs ≥1 year old (38.5% [50/130]) (P = 0.02). Group A (n = 35) exhibited median EBC of 1.05 × 109/L, and an eosinophilia ratio of 34.3% that was significantly higher (P < 0.05) than Group C (0.45 × 109/L and 4.1%, respectively). Both variables did not differ in Group B (n = 20), D (n = 39) or E (n = 36) in comparison to Group C (P > 0.05). By ROC curve analysis, only Group A generated a significant area under the curve (0.72). With EBC cutoff of 0.85 × 109 eosinophils/L, sensitivity and specificity were 65.7% and 70.8%, respectively. Eosinophil counts alone may be helpful in raising suspicion of an Ancylostoma spp. infection if further intestinal parasites and concurrent disorders are absent. However, local prevalence data and epidemiological findings should also be evaluated, since eosinophilia is less frequently observed with Ancylostoma spp. infections in the presence of comorbidities.

Accuracy of differences in blood and peritoneal glucose to differentiate between septic and non-septic peritonitis in horses.

Difference in blood and peritoneal glucose (DBPG) is used in clinical practice to support a diagnosis of septic peritonitis in horses. It is inexpensive, easy and rapid to perform. The aim of this study was to evaluate the accuracy of the DBPG to differentiate between septic and non-septic peritonitis in horses. Blood and peritoneal fluids were harvested from suspected animals. Plasma and peritoneal glucose levels, total nucleated cell count, direct microscopic and microbiological examinations of the peritoneal fluid were evaluated. Using DBPG levels, the animals were classified into two groups: difference ≥ 50 mg/dL (positive test) and difference < 50 mg/dL (negative test). Positive microbiological examination and/or presence of bacteria in direct microscopic examination was used as a gold standard to detect septic peritonitis. The accuracy parameters analysed were: sensitivity, specificity, and positive/negative predictive values, for which the results were respectively: 0.23, 0.91, 0.60 and 0.67. Due to poor accuracy, other cut-off margins and peritoneal glucose concentrations were evaluated. The test was considered most accurate when the DBPG was zero with sensitivity, specificity, and positive/negative predictive values of 0.85, 0.82, 0.73, 0.90 respectively. Peritoneal glucose concentrations alone were not a reliable feature to detect peritonitis. DBPG ≥50 mg/dL, widely used for the diagnosis of septic peritonitis, does not have a good accuracy and the DBPG = 0 has a better accuracy for detecting the disease.

Efficacy of cefquinome and a combination of cloxacillin and ampicillin for treatment of dairy cows with Streptococcus agalactiae subclinical mastitis.

A randomized clinical trial was conducted to assess efficacy of intramammary cloxacillin and ampicillin (CLOXIMM), intramammary cefquinome (CEFIMM), and intramuscular cefquinome (CEFIM) to treat Streptococcus agalactiae intramammary infections (Trial 1). Subsequently, two treatment groups were extended to assess whether CLOXIMM was not inferior to CEFIMM (Trial 2). Nine farms were included in the study. Milk samples were collected from all quarters of all lactating cows for microbiological identification of S. agalactiae. Positive cows were randomly allocated into four groups: CLOXIMM, CEFIMM, CEFIM, or negative control (CONTROL). Study outcomes were bacteriological cure at 14 (CURE14), 21 (CURE21), and 14 and 21 (CURE1421) days after treatment onset, and somatic cell count. Logistic regression was used to estimate the odds of cure between each treatment and CONTROL. Non-inferiority analysis was performed considering a one-sided 95% confidence interval (CI) and non-inferiority margins (Δ) of 0.10, 0.15, 0.20, and 0.25. Adjusted S. agalactiae bacteriological cure for CLOXIMM, CEFIMM, CEFIM, and CONTROL was 86, 98, 55, and 25% at day 14; 82, 93, 52, and 0% at day 21; and 82, 92, 40, and 0% at days 14 and 21, respectively. Treatment with CLOXIMM and CEFIMM resulted in greater bacteriological cure rates, as compared with CEFIM or CONTROL, which does not justify the use of CEFIM in S. agalactiae eradication programs. The CURE14 difference between CEFIMM and CLOXIMM was of 12.1 percentage points (95% CI: 0.056-0.184). CLOXIMM was considered not inferior to CEFIMM for Δ = 0.20 or 0.25 and inconclusive for Δ = 0.10 or 0.15. Thus, it should be pondered by veterinarians whether an expected 12.1 (5.6-18.4) percentage points increase in cure rate would justify the use of a fourth-generation cephalosporin, as opposed to a combination of traditional IMM drugs (cloxacillin and ampicillin) to treat S. agalactiae subclinical mastitis.

Brucellosis Prevalence in Brazilian Slaughterhouses with Different Meat Inspection Systems.

In Brazil, meat inspection occurs in a decentralized manner and consists of three types: (i) federal inspection (SIF), (ii) state inspection (SISP), and (iii) municipal inspection (SIM). The objective of this work was to discuss the three current inspection systems through the apparent prevalence of bovine brucellosis, a zoonosis that has an eradication program implemented by the Brazilian government. Nine abattoirs from federal, state, and municipal inspection systems were assessed and 1,490 animals were sampled. Serology for brucellosis was determined by the rose bengal test and the complement fixation test. The overall apparent prevalence (and 95% confidence interval) of brucellosis was 2.2% (1.5 to 2.9%). Apparent prevalence stratified by inspection system for SIF, SISP, and SIM was 0.4% (0.0 to 0.9%), 2.0% (0.8 to 3.2%), and 4.3% (2.5 to 6.1%), respectively. Multivariable logistic regression analysis revealed the odds ratio for finding an animal positive for brucellosis among inspection systems. A statistical difference ( P < 0.0015) was observed among surveillance systems, with SISP × SIF, SIM × SISP, and SIM × SIF having an odds ratio of 4,996, 2,304, and 11,494, respectively. Hence, the need for increasing official surveillance in state and municipal inspection systems seems to be necessary and could assist in the surveillance of bovine brucellosis and other diseases of interest to the federation. In addition, an increase in official presence would help to improve the selection of slaughtered cattle during ante- and postmortem inspection, with consequent impact on food safety and public health.

Do Milk Samples Stored for 12 Days after Collection Exhibit a Change in Composition Related to the Initial Bacterial Load?

Total bacterial count (TBC) is a tool used to assess milk quality and is associated with not only the initial sample contamination but also the sample storage time and temperature. Several countries have reported milk samples with a high TBC, and the influence of TBC on milk preservation remains unclear. Thus, the aim of this study was to evaluate the impact of the initial bacterial contamination level on the macrocomponents and somatic cell count (SCC) of raw milk samples preserved with bronopol and maintained at two storage temperatures (7 and 25°C) for up to 12 days. Thus, we collected milk samples from 51 dairy farms, which were divided into two groups according to the initial bacterial load: low TBC (<100,000 CFU/ml) and high TBC (≥100,000 CFU/ml). We analyzed the sample composition for protein, fat, total solids, lactose, milk urea nitrogen, and the SCC. We did not observe an effect from TBC and storage time and temperature on the concentration of protein, fat, total solids, and lactose. SCC changes were not observed for samples maintained under refrigeration (7°C); however, samples maintained at room temperature (25°C) exhibited a decrease in the SCC beginning on day 6 of storage. For milk urea nitrogen, values increased when the samples were maintained at room temperature, beginning on the ninth storage day. Samples with the preservative bronopol added and maintained under refrigeration may be analyzed up to 12 days after collection, regardless of the milk microbial load.

Streptococcus suis in employees and the environment of swine slaughterhouses in São Paulo, Brazil: Occurrence, risk factors, serotype distribution, and antimicrobial susceptibility.

Streptococcus suis is an important pathogen in the swine industry. This article is the first to report the occurrence, risk factors, serotype distribution, and antimicrobial susceptibility of S. suis recovered from employees and environmental samples of swine slaughterhouses in Brazil. Tonsillar swabs from all 139 pig-slaughtering employees and 261 environmental swabs were collected for detection of S. suis and serotyping by monoplex and multiplex polymerase chain reaction, respectively. Antimicrobial susceptibility was determined by the disk-diffusion method. Although S. suis was not detected in any of the tested employees, it was isolated from 25% of the environmental samples. Significant differences (P < 0.05) in the occurrence of S. suis were observed between slaughterhouses and between areas of low, medium, and high risk. The most frequent serotypes were 4 and 29, each accounting for 12% of the isolates, followed by 5, 12, 21, and 31, each accounting for 6%. High rates of susceptibility to the antimicrobials doxycycline (100%), ceftiofur (94%), ampicillin (81%), and cephalexin (75%) were observed. However, multidrug resistance was observed in all the isolates. Because S. suis is present in the environment of swine slaughterhouses, on carcasses and knives, as well as on the hands of employees in all areas, all employees are at risk of infection.

Streptococcus suis est un agent pathogène important dans l'industrie porcine. Cet article est le premier à rapporter la fréquence, les facteurs de risque, la distribution des sérotypes, et la sensibilité aux antimicrobiens d'isolats de S. suis provenant des employés et d'échantillons de l'environnement d'abattoirs de porcs au Brésil. Des échantillons des amygdales des 139 employés et 261 échantillons environnementaux furent prélevés pour détection de S. suis et sérotypage par réaction d'amplification en chaine monoplex et multiplex, respectivement. La sensibilité aux antimicrobiens a été déterminée par la méthode de diffusion en disque. Bien que S. suis ne fut isolé d'aucun des employés testés, la bactérie a été isolée de 25 % des échantillons environnementaux. Des différences significatives (P < 0,05) dans la fréquence de S. suis furent observées entre les abattoirs et entre les zones à risque faible, moyen, et élevé. Les sérotypes 4 et 29 étaient les plus fréquents, comptant chacun pour 12 % des isolats, suivi des sérotypes 5, 12, 21, et 31, chacun comptant pour 6 %. Des pourcentages élevés de sensibilité à la doxycycline (100 %), au ceftiofur (94 %), à l'ampicilline (81 %) et à la céphalexine (75 %) ont été notés. Toutefois, de la résistance multiple fut observée chez tous les isolats. Étant donné que S. suis est présent dans l'environnement des abattoirs de porc, sur les carcasses et les couteaux, ainsi que sur les mains des employés dans toutes les zones, tous les employés sont à risque de s'infecter.(Traduit par Docteur Serge Messier).

Evaluation of short-interfering RNAs treatment in experimental rabies due to wild-type virus.

We have evaluated the efficacy of short-interfering RNAs targeting the nucleoprotein gene and also the brain immune response in treated and non-treated infected mice. Mice were inoculated with wild-type virus, classified as dog (hv2) or vampire bat (hv3) variants and both groups were treated or left as controls. No difference was observed in the lethality rate between treated and non-treated groups, although clinical evaluation of hv2 infected mice showed differences in the severity of clinical disease (p=0.0006). Evaluation of brain immune response 5 days post-inoculation in treated hv2 group showed no difference among the analyzed genes, whereas after 10 days post-inoculation there was increased expression of 2',5'-oligoadenylate synthetase 1, tumor necrosis factor alpha, interleukin 12, interferon gamma, and C-X-C motif chemokine 10 associated with higher expression of N gene in the same period (p<0.0001). In hv2 non-treated group only higher interferon beta expression was found at day 5. The observed differences in results of the immune response genes between treated and non-treated groups is not promising as they had neither impact on mortality nor even a reduction in the expression of N gene in siRNA treated animals. This finding suggests that the use of pre-designed siRNA alone may not be useful in rabies treatment.