Antibacterial spectrum and cytotoxic activities of serrulatane compounds from the Australian medicinal plant Eremophila neglecta.

AIMS: To determine the antibacterial spectrum and cytotoxic activities of serrulatane compounds from the Australian plant Eremophila neglecta. METHODS AND RESULTS: Antimicrobial activities of serrulatane compounds 8,19-dihydroxyserrulat-14-ene (1) and 8-hydroxyserrulat-14-en-19-oic acid (2) were tested against Gram-negative and Gram-positive bacteria including human and veterinary pathogens and some multidrug-resistant isolates. Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of the compounds were determined by broth microdilution assay. Both compounds exhibited antibacterial activity against all Gram-positive test strains. They showed antimycobacterial activity against isolates of Mycobacterium fortuitum and Mycobacterium chelonae. Of the five Gram-negative bacteria tested, only Moraxella catarrhalis showed susceptibility to the compounds. Cytotoxic activities were tested in the Vero cell line. Compound 1 showed more activity than 2 in both antibacterial and cytotoxicity assays with cytotoxicity at concentrations similar to the MBC. CONCLUSIONS: Serrulatane compounds showed significant activity against medically important bacteria, with 1 exhibiting stronger antibacterial activity. However, they also displayed toxicity to mammalian cells. SIGNIFICANCE AND IMPACT OF THE STUDY: Serrulatanes are of interest as novel antibacterial compounds for use in biomedical applications; this study reports data obtained with a range of bacterial strains and mammalian cells, essential for assessing the capabilities and limitations of potential applicability of these compounds.

Bacteriophage-mediated transduction of antibiotic resistance in enterococci.

AIMS: Temperate bacteriophages are bacterial viruses that transfer genetic information between bacteria. This phenomenon is known as transduction, and it is important in acquisition of bacterial virulence genes and antimicrobial resistance determinants. The aim of this study was to demonstrate the role of bacteriophages in gene transfer (antibiotic resistance) in enterococci. METHODS AND RESULTS: Three bacteriophages from environmental samples isolated on pig host strains of Enterococcus gallinarum and Enterococcus faecalis were evaluated in transduction experiments. Antibiotic resistance was transferred from Ent. gallinarum to Ent. faecalis (tetracycline resistance) and from Ent. faecalis to Enterococcus faecium, Enterococcus hirae/durans and Enterococcus casseliflavus (gentamicin resistance). CONCLUSIONS: Bacteriophages play a role in transfer of antibiotic resistance determinants in enterococci. SIGNIFICANCE AND IMPACT OF THE STUDY: This study confirms previous suggestions on transduction in enterococci, in particular on interspecies transduction. Interspecies transduction is significant because it widens the range of recipients involved in antimicrobial resistance transfer.

Incidence of class 1 integron and other antibiotic resistance determinants in Aeromonas spp. from rainbow trout farms in Australia.

There is limited information on antibiotic resistance determinants present in bacteria of aquaculture origin in Australia. The presence of integron and other resistance determinants was investigated in 90 Aeromonas isolates derived from nine freshwater trout farms in Victoria (Australia). Polymerase chain reaction was carried out for the detection of integrase genes Int1, Int2 and Int3, gene cassette array, integron-associated aadA, sul1 and qac1 genes, streptomycin resistance genes strA-strB, ß-lactamase resistance genes bla(TEM) and bla(SHV) , and tetracycline resistance genes tetA-E and tetM. Clonal analysis was performed by pulsed-field gel electrophoresis (PFGE). Class 1 integrons were detected in 28/90 (31%) and class 2 and class 3 in none of the strains, aadA gene in 19/27 (70%) streptomycin-resistant strains, sul1 in 13/15 (86.7%) sulphonamide-resistant strains and qac1 gene in 8/28 (28.6%) integron-bearing strains. Five strains from two different farms carried gene cassettes of 1000 bp each containing the aadA2 gene and PFGE analysis revealed genetic relatedness. tetC was detected in all and tetA in 9/18 (50%) tetracycline-resistant strains. The strA-strB, bla(TEM) or bla(SHV) genes were not detected in any of the strains. Aeromonas spp. carrying integrons and other resistance genes are present in farm-raised fish and sediments even though no antibiotics were licensed for use in Australian aquaculture at the time of the study.

Antimicrobial use in the Australian pig industry: results of a national survey.

OBJECTIVE: To describe how various antimicrobials are used in commercial pig herds in Australia and for what disease conditions. PROCEDURE: Managers of large pig herds (> 200 sows) across Australia and their veterinarians participated in an internet-based survey in 2006. Questions were asked about herd management, the occurrence of bacterial diseases and the type and frequency of antimicrobial use. An antimicrobial usage index for each herd was derived as a summary of the risk of selection for antimicrobial resistance. Relationships between responses were explored with univariate and multivariate analysis. RESULTS: Responses were received for 197 herds estimated to represent at least 51% of all large pig herds in Australia. Most piggeries relied on drugs of low importance in human medicine (e.g. tetracyclines, penicillins and sulfonamides). For the two drugs of high importance in human medicine that can be legally prescribed to pigs in Australia, ceftiofur use was reported in 25% of herds and virginiamycin in none. Infections attributed to Lawsonia, Mycoplasma and Escherichia coli motivated the most use of antimicrobials. No useful association was found between management factors and the antimicrobial use index. CONCLUSION: Most antimicrobial use in the Australian pig industry is based on drugs of low importance to public health. Enhanced control of E. coli infections without reliance on antimicrobials would further reduce the risk of selecting for antimicrobial resistance relevant to public health. The amount of variation in the usage index between herds suggests that antimicrobial use should be constantly reviewed on a herd by herd basis.

Rhodococcus equi (Prescottella equi) vaccines; the future of vaccine development.

For decades researchers have been targeting prevention of Rhodococcus equi (Rhodococcus hoagui/Prescottella equi) by vaccination and the horse breeding industry has supported the ongoing efforts by researchers to develop a safe and cost effective vaccine to prevent disease in foals. Traditional vaccines including live, killed and attenuated (physical and chemical) vaccines have proved to be ineffective and more modern molecular-based vaccines including the DNA plasmid, genetically attenuated and subunit vaccines have provided inadequate protection of foals. Newer, bacterial vector vaccines have recently shown promise for R. equi in the mouse model. This article describes the findings of key research in R. equi vaccine development and looks at alternative methods that may potentially be utilised.

Ecology of Rhodococcus equi.

A selective broth enrichment technique was used to study the distribution of Rhodococcus equi in soil and grazing animals. Rhodococcus equi was isolated from 54% of soils examined and from the gut contents, rectal faeces and dung of all grazing herbivorous species examined. Rhodococcus equi was not isolated from the faeces or dung of penned animals which did not have access to grazing. The isolation rate from dung was much higher than from other samples and this was found to be due to the ability of R. equi to multiply more readily in dung. Delayed hypersensitivity tests were carried out on horses, sheep and cattle, but only horses reacted significantly. The physiological characteristics of R. equi and the nature of its distribution in the environment suggested that R. equi is a soil organism.

Antibiotic use in animal feed and its impact on human healt.

Antibiotic resistance in bacteria that cause disease in man is an issue of major concern. Although misuse of antibiotics in human medicine is the principal cause of the problem, antibiotic-resistant bacteria originating in animals are contributory factors, with some types of resistance in some species of bacteria. Antibiotics are added to animal feeds to treat and prevent infections and to improve growth and production. Until recently, the major concerns about incorporation of antibiotics in animal feeds related to antibiotic residues in products from treated animals. Although, in 1969, the Swann (1969) report drew attention to the potential for antibiotic-resistant bacteria to spread from treated animals via the food chain, there was little response until the detection of vancomycin-resistant enterococci in animals fed a related glycopeptide, avoparcin. Subsequently, attention started to focus on the issue and other examples of transfer of resistant bacteria through the food chain, such as enterococci resistant to quinupristin-dalfopristin or to everninomicin, fluoroquinolone-resistant campylobacters and multiresistant Escherichia coli, and salmonella such as Salmonella typhimurium DT104. Reviews and committees in many countries have highlighted the need for better control of licensing of antibiotics, and codes for prudent use of antibiotics by veterinary practitioners and farmers. The continued use of antibiotic growth promoters has been questioned and there is a need to ensure that antibiotics important in human medicine are not used therapeutically or prophylactically in animals.

Opening and closing mechanisms of the larynx.

Opening and closing of the larynx are determined by the intrinsic and extrinsic muscles acting on the elastic forces in the tongue, pharynx, larynx, and trachea. The pharynx is opened or closed by two mechanisms: (1) Contractions of the cricothyroid and of the intrinsic muscles of the larynx open and close the vocal cords. (2) The false cords, ventricle, and true cords accordion open or close in a bellows mechanism. We conclude that the posterior cricoarytenoid opens the laryngeal airway. The cricothyroid together with the posterior cricoarytenoid accentuates this opening. The larynx is also opened by the geniohyoid, mylohyoid, sternothyroid, and middle constrictor. The thyrohyoid, cricothyroid, sternohyoid, and inferior constrictor close the laryngeal airway. Abnormalities in the soft tissues of the neck or of the innervation of the larynx, pharynx, and neck muscles may severely interfere with patency of the laryngeal airway. This occurs in such conditions as vocal cord paralysis, sleep apnea, multiple sclerosis, amyotrophic lateral sclerosis, spastic dysphonia, mandibular fractures or hypodevelopment, and cerebrovascular disease.

Outpatient surgery and anesthesia.

Outpatient surgery and anesthesia are offered as a means of reducing the costs of health care delivery. Certain principles for outpatient anesthesia are apparent. Inhalational anesthesia is preferred over many intravenous agents. If intravenous agents are used, the doses must be limited. Intravenous agents that are quickly metabolized or redistributed in the body away from the brain are preferred. Inhalational anesthesia inductions that avoid or minimize intravenous agents are preferred. Nerve blocks are useful for outpatient surgery, but may require supplementation with intravenous agents, which in turn may slow recovery.

Studies of the pathogenesis of Rhodococcus equi infection in foals.

Pyogranulomatous pneumonia was induced in Thoroughbred foals by intranasal challenge with freeze-dried cultures of Rhodococcus equi (previously Corynebacterium equi). The incubation period was about 18 days and clinical signs were not seen for a further week. There were marked seasonal and individual foal differences in responses to infection. Elevations in serum caeruloplasmin oxidase activity and copper concentrations appeared to be sensitive indicators of infection. Serum zinc concentrations and serum alpha-mannosidase and alkaline phosphatase activities fell in the more severely infected foals. Use of trace elements and trace element-related parameters along with faecal culture for R. equi could prove useful for early diagnosis of field cases.

Campylobacter species in cats and dogs in South Australia.

BACKGROUND: Campylobacter enteritis was the most frequently notified infectious disease in Australia in 1996 and Campylobacter species have been associated with extra-intestinal infections such as purulent arthritis and Guillian-Barré syndrome. Dogs and cats are known to carry campylobacteria and contact with household pets have been implicated as possible sources of human infection. OBJECTIVE: To provide information on the species of campylobacter carried by cats and dogs in South Australia. METHODS: Faecal samples were collected from stray and owned cats and dogs and feral cats. Campylobacter-like organisms were isolated using selective media and filtration methods. They were then characterised by biochemical tests, antibiotic resistance and growth patterns under various conditions. Husbandry factors that could have influenced the carriage rates were examined both as single variables and in a multivariate logistic regression. RESULTS: Campylobacter upsaliensis and C jejuni were found in 11% and 4% of cats, respectively, whereas 34% dogs carried C upsaliensis, 7% C jejuni and 2% C coli. Intensive housing and open drains were found to be significant risk factors and increased the carriage rate by 2 and 2.6 times, respectively. CONCLUSION: Dogs and cats are a potential reservoir for human enteric infections with campylobacters.

Seroprevalence to Leptospira interrogans serovar hardjo in merino stud rams in South Australia.

A serological survey of 2160 Merino stud rams on 36 farms detected positive reactions greater than or equal to 1/100 in 42% of animals using the microscopic agglutination test (MAT) to Leptospira interrogans serovar hardjo. Twenty flocks had seroprevalence values greater than 30% with 15 flocks having values > or = 60%. The enzyme-linked immunosorbent assays showed that 47% and 3% of rams on the 36 farms were positive for IgG and IgM antibodies, respectively. Forty-five percent of hardjo reactions were in rams that had not been exposed to cattle. Significant correlations were found between IgM reactors and creek/dam water pumped into troughs, and between MAT/IgG reactors and total flock size. No statistical relationships were detected between positive reactors and two different annual average rainfall gradients, the time of the year in which samples were obtained, or agricultural regions of South Australia. Infections with an organism of the Sejroe serogroup is widespread in Merino stud rams.

An outbreak of Mycobacterium bovis infection in cats in an animal house.

An outbreak of tuberculosis due to Mycobacterium bovis in cats in an animal house was investigated. It was concluded that the index case was infected by ingestion of contaminated meat obtained from a knackery and that some of the other cases were infected by inhalation of tubercle bacilli shed from a discharging sinus in the index case. A possum was also infected and a research worker apparently received a significant challenge.

Methicillin-resistant Staphylococcus aureus in a population of horses in Australia.

OBJECTIVE: To evaluate if methicillin-resistant Staphylococcus aureus (MRSA) is present in the horse population in Australia. DESIGN: A two-part retrospective study of laboratory submissions of microbial culture results from horses. METHODS: Part A: medical records of 216 horses that had MRSA screening performed on nasal swabs collected over a 30-day period at admission to the Scone Equine Hospital Clovelly Intensive Care Unit were retrieved. Part B: laboratory records from 2004 to 2009 of culture submissions to the Scone Veterinary Laboratory were reviewed and cultures that grew MRSA were identified. The MRSA isolates from Parts A and B were genotyped over an 18-month period. RESULTS: MRSA screening of 216 horses identified eight (3.7%) positive samples. MRSA was isolated from cultures of 80 (0.002%) clinical bacteriology samples over a 6-year period. Genotypic analysis was performed on 36 isolates. All MRSA characterised had the same pulse field gel electrophoresis pattern (type 1), with eight closely related subtypes identified (subtypes A-F and H) and 66% of isolates classified as subtype D, which multilocus sequence and staphylococcal cassette chromosome mec typing analysis identified as ST612-MRSA-IVa, a clonal complex (CC) 8 S. aureus strain. Antimicrobial resistance to more than two classes of antimicrobials was common. CONCLUSIONS: MRSA was present in a population of horses in Australia. Genotypic analysis of the isolates identified the MRSA strain as CC8 S. aureus. Further research needs to be undertaken to evaluate MRSA infection and colonisation of horses and personnel in Australia.

Carriage of methicillin-resistant Staphylococcus aureus by veterinarians in Australia.

OBJECTIVE: To estimate the prevalence of carriage of methicillin-resistant Staphylococcus aureus (MRSA) among Australian veterinarians. METHODS: Individuals attending veterinary conferences in Australia in 2009 were recruited to provide nasal swabs and complete a questionnaire about their professional activities. Swabs were processed by standard methods for detecting MRSA and questionnaire responses were used to group veterinarians according to their areas of major work emphasis (species and practice type). Prevalence was estimated for each of these grouping and contingency tables and regression tree analysis used to explain the variation in MRSA carriage. RESULTS: Among the 771 respondents 'industry and government veterinarians' (controls) had the lowest prevalence of MRSA carriage at 0.9%. Veterinarians with horses as a major area of work emphasis had a prevalence of 11.8% (13-fold that of controls) and those whose only major emphasis was horses had a prevalence of 21.4% (23-fold that of controls). Veterinarians with dogs and cats as a major activity had a 4.9% prevalence (5-fold that of controls). Prevalence rates for other major activities (pigs, dairy and beef cattle, avian and wildlife) were also increased, but were estimated from smaller numbers of respondents. Regression tree analysis clearly isolated equine veterinarians and dog and cat practitioners as groups at increased risk of carriage of MRSA. CONCLUSION: Carriage of MRSA is a notable occupational health issue for veterinarians in clinical practice in Australia, particularly those who work with horses.

Antimicrobial resistance and virulence gene profiles in multi-drug resistant enterotoxigenic Escherichia coli isolated from pigs with post-weaning diarrhoea.

This study aimed to characterize antimicrobial resistance and virulence genes in multi-drug resistant enterotoxigenic Escherichia coli (ETEC) isolates (n=117) collected from porcine post-weaning diarrhoea cases in Australia (1999-2005). Isolates were serotyped, antibiogram-phenotyped for 12 antimicrobial agents and genotyped by PCR for 30 plasmid-mediated antimicrobial resistance genes (ARGs), 22 intestinal and 38 extraintestinal E. coli virulence genes (VGs). Nine serogroups were identified, the most prevalent being O149 (46.2%), O141 (11.2%) and Ont (31.6%). None of the isolates showed resistance to ceftiofur or enrofloxacin and 9.4% were resistant to florfenicol. No corresponding extended-spectrum/AmpC ß-lactamase, fluoroquinolone or floR ARGs were detected. An antimicrobial resistance index (ARI) was calculated from the combined data with a weighting for each antimicrobial agent dependent upon its significance to human health. Serogroup O141 isolates had a significantly higher ARI due to an elevated prevalence of aminoglycoside ARGs and possession of more virulence genes (VGs), including ExPEC or EHEC adhesins (bmaE, sfa/focDE, fimH, ihA) in toxin-producing strains that lacked the normally associated F4 and F18 fimbriae. Few associations between ARGs and VGs were apparent, apart from tetC, sfa/focDE and ompT which, for a sub-set of O141 isolates, suggest possible plasmid acquisition from ExPEC. The multi-drug resistant ETEC ARG/VG profiles indicate a high probability of considerable strain and plasmid diversity, reflecting various selection pressures at the individual farm level rather than emergence and lateral spread of MDR resistant/virulent clones.