9 Barton BM, Harding GP, Zuccarelli AJ. A general method for detecting and sizing large plasmids.

BackgroundAmong coagulase-negative staphylococci, Staphylococcus haemolyticus is the second most frequently isolated species from human blood cultures and has the highest level of antimicrobial resistance. This species has zoonotic character and is prevalent both in humans and animals. Recent studies have indicated that methicillin-resistant S. haemolyticus (MRSH) is one of the most frequent isolated Staphylococcus species among neonates in intensive care units. The aim of this study was to determine the presence of MRSH in different groups of companion animals and to characterize isolates according their antimicrobial resistance.MethodsSamples (n = 754) were collected from healthy and diseased dogs and cats, female dogs in pure-breed kennels, healthy horses, and kennel owners. Classical microbiological tests along with molecular testing including PCR and 16S rRNA sequencing were performed to identify MRSH. Clonality of the isolates was assessed by Pulsed Field Gel Electrophoresis using the SmaI restriction enzyme. Antimicrobial susceptibility testing was performed using the broth micro-dilution method. Detection of genes encoding antimicrobial resistance was performed by PCR. Statistical analysis was performed using the R Project of Statistical Computing, “R 1.8.1” package.ResultsFrom a total of 754 samples tested, 12 MRSH isolates were obtained. No MRSH were found in horses and cats. Eleven isolates were obtained from dogs and one from a kennel owner. Ten of the dog isolates were detected in pure-breed kennels. The isolates demonstrated the same clonality only within separate kennels.The most frequent resistances of MRSH isolates was demonstrated to benzylpenicillin (91.7%), erythromycin (91.7%), gentamicin (75.0%), tetracycline (66.7%), fluoroquinolones (41.7%) and co-trimoxazole (41.7%). One isolate was resistant to streptogramins. All isolates were susceptible to daptomycin, rifampin, linezolid and vancomycin. The clone isolated from the kennel owner and one of the dogs was resistant to beta-lactams, macrolides, gentamicin and tetracycline.ConclusionsPure-breed kennels keeping 6 or more females were determined to be a risk factor for the presence of MRSH strains. MRSH isolated from companion animals were frequently resistant to some classes of critically important antimicrobials, although they remain susceptible to antibiotics used exclusively in human medicine.

The aim of this study was to characterize Staphylococcus pseudintermedius for its antimicrobial resistance and virulence factors with a special focus on methicillin-resistant (MRSP) strains isolated from sick dogs in Lithuania. Clinically sick adult dogs suffering from infections (n=214) and bitches with reproductive disorders (n=36) from kennels were selected for the study. Samples (n=192) from the 250 tested (76.8%) dogs were positive for Staphylococcus spp. Molecular profiling using the species-specific nuc gene identified 51 isolates as S. pseudintermedius (26.6% from a total number of isolated staphylococci) of which 15 isolates were identified as MRSP. Ten MRSP isolates were isolated from bitches with reproductive disorders from two large breeding kennels. Data on susceptibility of S. pseudintermedius to different antimicrobials revealed that all isolates were susceptible to vancomycin, daptomycin and linezolid. Two isolates (3.9%) were resistant to rifampicin. A high resistance was seen towards penicillin G (94.1%), tetracycline (64.7%) and macrolides (68.7%). Resistance to fluoroquinolones ranged from 25.5% (gatifloxacin) to 31.4% (ciprofloxacin). The most prevalent genes encoding resistance included blaZ, aac(6')- mecA, and tet(M). The Luk-I gene encoding a leukotoxin was detected in 29% of the isolates, whereas the siet gene encoding exfoliative toxin was detected in 69% of the S. pseudintermedius isolates. This report of MRSP in companion animals represents a major challenge for veterinarians in terms of antibiotic therapy and is a concern for both animal and public health.

The aim of the study was to isolate cultivable gut microbiota from European pond turtles kept at the Lithuanian Zoo and to determine antimicrobial resistance of the isolates. The study subjects included 8 elderly turtles living at the Lithuanian Zoo for about 50 years as well as their offspring -24 young individuals (1-2 years old) that were hatched at the same zoo. Animals were not exposed by treatment with antimicrobials during the last 3 years.Gut samples were taken from the cloaca and inoculated onto universal media. Isolates then were identified using sequence analysis of 16S rRNA.The antimicrobial susceptibility testing was performed using the agar diffusion method according to Kirby-Bauer. Clinical breakpoints according to CLSI whenever possible, were used for interpretation of susceptibility. Bacterial isolates resistant to at least three antimicrobials of different classes were treated as multi-resistant.Fifty-two bacterial isolates were obtained and identified from turtle gut samples. The most prevalent genera included Aeromonas, Chryseobacterium and Citrobacter. Fifty percent of the isolates obtained from elderly turtles (CI 95% -19.01-80.99) and 54.8% (CI 95% -39.75-69.85) of the isolates from young animals were identified as multi-resistant. The most common resistance rates of the isolates from both groups of the turtles were observed toward ampicillin (86.6%), ciprofloxacin (61.5%) and gentamicin (40.4%). The lowest number of resistant isolates were detected toward combination of sulfamethoxazole-trimethoprim (26.9%). The study revealed that European pond turtles kept in captivity are carriers of multi-resistant bacteria however, further studies need to be performed to investigate whether the resistant microorganisms are natural microbiota for this species or they were acquired in the zoo.

ABSTRACT:Here, the ability of Pediococcus pentosaceus and Pediococcus acidilactici to utilise potato tuber juice for cell synthesis without an external nutrient supplement was investigated, and the influence of lactic acid bacteria (LAB) grown in this substrate on the growth performance of new-born calves, as well as blood biochemical and faecal microbiological parameters was evaluated. Calves were selected based on the analogy principle, treatment group (n = 21), control group (n = 27). Calves in the treatment group were administered 50 ml of fermented potato tubers juice containing 9.6 log CFU/ml of LAB mixture for 14 days. Also, determination of antimicrobial activities of tested LAB against a variety of pathogenic and opportunistic bacterial strains previously isolated from diseased cattle was performed. It was found that LAB supernatants effectively inhibited the growth of Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, Salmonella enterica, Corynebacter spp., Klebsiella pneomoniae, Enterococcus faecalis, and Bacillus cereus (the diameters of the inhibition zones varied between 11.0 ± 0.3 mm and 17.0 ± 0.6 mm). Thus, potato juice can be used as an alternative substrate for LAB cultivation (LAB cell concentration 9.6 ± 0.07 log CFU/ml). After lyophilisation (-48 °C) and spray-drying (+150 °C) viable cell concentrations in the fermented potato juice powder were 9.18 ± 0.09 log CFU/g and 9.04 ± 0.07 log CFU/g, respectively. The 50 ml of fermented potato tuber juice containing 9.6 log CFU/ml of LAB, administered every day for 14 days, reduced the risk of developing acidosis (stabilised blood pH; P < 0.05), reduced lactates and PCO 2 concentration (P < 0.05) and the risk of liver lesions (reduced serum alanine aminotransferase concentration; P < 0.005) in blood and E. coli in the faeces of new-born calves.