A zoospore inhibition technique to evaluate the activity of antifungal compounds against Batrachochytrium dendrobatidis and unsuccessful treatment of experimentally infected green tree frogs (Litoria caerulea) by fluconazole and benzalkonium chloride

Effective and safe treatments of chytridiomycosis in amphibians, caused by Batrachochytrium dendrobatidis, are needed to help prevent mortality in captive programs for threatened species, to reduce the risk of spread, and to better manage the disease in threatened populations. We describe a simple method to determine minimum inhibitory concentrations (MICs) of antifungal agents that involves adding zoospores to various drug concentrations in 96 well plates and microscopic observation after four days. We report results from testing 10 commercially available antifungal compounds: benzalkonium chloride (<0.78 μg/ml), povidone iodine (312.5 μg/ml), amphotericin B (3.125 μg/ml), fluconazole (<1.56 μg/ml), itraconazole (<1.56 μg/ml), enilconazole (<1.56 μg/ml), mercurochrome (6.25 μg/ml), sodium chloride (12.5 mg/ml), methylene blue (<1.56 μg/ml) and Virkon (3.125 μg/ml). For treatment trials of juvenile Litoria caerulea, baths of benzalkonium chloride at 1 mg/L and fluconazole at 25 mg/L were used on 18 experimentally infected frogs per treatment. Although these treatments resulted in longer survival times (mean 43.7 ± 11.3 days) than in the untreated controls (37.9 ± 9.3 days), the mortality rate was still 100%. Higher doses of fluconazole are suggested for further animal trials.     

Bacterial flora on Cascades frogs in the Klamath mountains of California

Amphibians are experiencing global declines due in part to the infectious disease chytridiomycosis. Some symbiotic bacteria residents on frog skin have been shown to inhibit the growth of Batrachochytrium dendrobatitis (Bd) but few studies have attempted to fully describe the resident bacterial flora of frog skin. We cultured and sequenced 130 bacterial isolates from frogs collected from the California Klamath Range, recovering predominantly Gram-negative bacteria from 20 higher order taxa and 31 genera. There were also a large number of unclassifiable isolates. Forty-three isolates were assessed for their ability to inhibit the growth of Bd in vitro; of these, two had strong and three had slight anti-Bd activity. We suggest that many bacterial species may play a secondary role in Bd resistance, acting synergistically with inhibitory species. Future research is required in order to characterize these interactions. Understanding the relationships between bacterial strains may be important in predicting and managing the effects of future anti-Bd treatments such as antimicrobial compounds or probiotic bacteria.     

猪大肠杆菌和葡萄球菌的分离鉴定及耐药性分析

对湘西北地区某猪场患病的仔猪的粪便及病变的脾脏、肝脏、淋巴结、肺脏、肾水肿液进行病原菌分离鉴定,并用Kirby-Bauer法测定分离菌株对15种抗菌药的敏感性。结果表明,患病仔猪分离到大肠杆菌和猪葡萄球菌两种细菌。其中大肠杆菌对庆大霉素、头孢呋肟、丁胺卡那霉素、四环素、先锋必素敏感,对氯霉素、氧哌嗪青霉素、先锋五号、头孢曲松钠、新霉素中介,对羧苄青霉素、强力霉素、青霉素、氨苄青霉素、红霉素有较高的耐药性。葡萄球菌对先锋霉素Ⅴ、丁胺卡那霉素敏感,对头孢曲松钠中介,对其他抗生素耐药。其中大肠杆菌对药物敏感率、中介率、耐药率均为33.33%;葡萄球菌的敏感率为13.33%,中介率为6.67%,耐药率为80.00%。     

Molecular survey of Dirofilaria immitis and Dirofilaria repens by direct PCR for wild caught mosquitoes in the Republic of Korea

Adult mosquito collections using New Jersey light traps and Black-hole light traps were conducted to determine the potential vectors and the relative mosquito infection rates of Dirofilaria immitis and Dirofilaria repens in Gyeonggi and Gangwon Provinces, Republic of Korea, 2005. Dirofilaria spp. were confirmed by polymerase chain reaction (PCR) using species-specific primers for D. immitis and D. repens. Minimum field infection rates (MFIR) [MFIR = (number infected pools/total number of mosquitoes) x 1000] of 12 pools/2059 total number mosquitoes (5.8) and 21 pools (10.1) of the wild caught mosquitoes were positive by PCR for D. immitis and D. repens, respectively. Dual infections of both D. immitis and D. repens were detected by PCR in eight pools (3.9). Anopheles sinensis sensu lato (includes An. sinensis s.s., An. pullus, An. kleini, An. belenrae, and An. lesteri), An. sineroides, Aedes vexans nipponi, Culex pipiens and Armigeres subalbatus were found to be infected and are potential vectors of D. immitis and D. repens in Korea. Infections observed in mosquitoes represent potential health risks for domestic animal and human populations.     

Perspectives on canine and feline hepatozoonosis

Two species of Hepatozoon are currently known to infect dogs and cause distinct diseases. Hepatozoon canis prevalent in Africa, Asia, southern Europe, South America and recently shown to be present also in the USA causes infection mainly of hemolymphoid organs, whereas Hepatozoon americanum prevalent in the southeastern USA causes myositis and severe lameness. H. americanum is transmitted by ingestion of the Gulf Coast tick Amblyomma maculatum and also by predation on infected prey. H. canis is transmitted by Rhipicephalus sanguineus, in South America also by Amblyomma ovale, and has also been shown to be transmitted transplacentally. Hepatozoonosis of domestic cats has been described mostly from the same areas where canine infection is present and the exact identity of the species which infect cats, their pathogenicity and vectors have not been elucidated. The diagnosis of hepatozoonosis is made by observation of gamonts in blood smears, histopathology, PCR or serology. The main treatment for H. canis is with imidocarb dipropionate whereas H. americanum infection is treated with an initial combination of trimethoprim-sulfadiazine, pyrimethamine and clindamycin followed by maintenance with decoquinate. Treatment for both diseases has not been reported to facilitate complete parasite elimination and new effective drugs are needed for the management of these infections. Prevention of hepatozoonosis should be based on avoidance of oral ingestion of infected tick vectors and infected prey.     

Antimicrobial susceptibility testing of Spanish field isolates of Brachyspira hyodysenteriae

This study is the first conducted in Spain to evaluate antimicrobial susceptibility of field isolates of Brachyspira hyodysenteriae. One hundred and eight isolates of the bacterium, recovered from different Spanish swine farms between 2000 and 2007, were investigated. The minimum inhibitory concentrations (MIC) of erythromycin, tylosin, tiamulin, valnemulin, clindamycin and lincomycin were determined using a broth microdilution technique. Most of the isolates showed poor susceptibility to erythromycin (MIC₉₀ > 256 μg/ml), tylosin (MIC₉₀ > 256 μg/ml), clindamycin (MIC₉₀ > 4 μg/ml) and lincomycin (MIC₉₀ = 128 μg/ml). Reduced susceptibility to tiamulin and valnemulin was observed with a MIC > 2 μg/ml in 17.6% and 7.41% of the B. hyodysenteriae isolates, respectively. Moreover, a survival analysis permitted the detection of an increasing trend in the MIC values for almost all the antimicrobials used in the treatment of swine dysentery when comparing recent isolates (from 2006 to 2007) with those recovered in earlier years (between 2000 and 2004).     

In vitro effects of four tropical plants on three life-cycle stages of the parasitic nematode, Haemonchus contortus

Alcoholic extracts of four tropical plants (Zanthoxylum zanthoxyloides, Newbouldia laevis, Morinda lucida and Carica papaya) were screened in vitro for potential anti-parasitic effects against eggs, infective larvae and adult Haemonchus contortus. Significant effects were obtained with all four plants but differences were observed depending on the parasitic stage. The effects of the four plant extracts were similar on egg hatching and were dose dependent. In contrast, no dose-response relationship was found for infective larvae and adult worms, although more potent effects were usually observed with the highest concentrations. Using a larval inhibition migration test, extracts of fagara (Z. zanthoxyloides) were found to be less active against Haemonchus infective larvae than were the other plants. N. laevis was found to be highly and rapidly effective against adult worms. Overall, these in vitro results suggest that these four plants, traditionally used by small farmers in Western Africa, do possess anti-parasitic properties. These effects remain to be confirmed through in vivo studies.     

Climate and the epidemiology of gastrointestinal nematode infections of sheep in Europe

The free-living stages of gastrointestinal nematode parasites of sheep are strongly affected by climate. Thus, extreme heat and cold are detrimental to development and survival, while, within tolerable limits, increasing temperatures generally accelerate development but increase mortality. Moisture is needed for development and translation of larvae from faeces to pasture, and so rainfall is a limiting factor for transmission. Together, these factors underpin seasonal patterns of infection in sheep, as well as geographic variation in the epidemiology and relative importance of different species within Europe. Local knowledge and experience enable treatment to be targeted appropriately to prevent dangerous levels of infection. This traditional know-how can be supplemented by predictive epidemiological models, built on thorough understanding of the influence of climate on larval availability. However, management also has a dominant role in determining patterns of infection, and is itself influenced by climate. Current geographic variation in nematode epidemiology across Europe, and knowledge of systems from outside Europe, can provide only limited perspectives on the likely effects of climate change on disease in future. This is because disease arises from complex interaction between host and parasite factors, and the implementation of optimal control strategies to meet new challenges will be slowed by the inertia of current systems. Approaches to nematode control must therefore take account not only of parasite biology, but also the forces that shape sheep farming systems and management decisions.     

乳酸菌―芽孢菌发酵黄芪对小鼠免疫功能作用的研究简

试验旨在探讨乳酸菌―芽孢菌发酵黄芪对小鼠免疫功能的影响。将84只小鼠分成6组,每组14只,分别为生理盐水组（阴性对照组）、未发酵黄芪组（阳性对照组,黄芪生药浓度为150 mg/mL）、试验Ⅰ组（黄芪生药浓度为60 mg/mL）、试验Ⅱ组（黄芪生药浓度为90 mg/mL）、试验Ⅲ组（黄芪生药浓度为120 mg/mL）、试验Ⅳ组（黄芪生药浓度为150 mg/mL）。小鼠采用灌胃方式给药30 d,最后处死小鼠测定其体重、免疫器官（胸腺和脾脏）系数、免疫因子（IgG、IL-6、sIgA）。结果显示,给药30 d后,试验组小鼠体重明显升高;试验Ⅰ组免疫器官系数极显著高于对照组（P＜0.01）;试验Ⅲ、Ⅳ组免疫因子IgG和sIgA均极显著高于对照组（P＜0.01）。由此可知,乳酸菌―芽孢菌发酵黄芪对小鼠的免疫功能有很好的促进作用。     

Diversity of Campylobacter jejuni and Campylobacter coli Genotypes from Human and Animal Sources from Rio de Janeiro, Brazil

To compare the genotypes of Campylobacter jejuni and Campylobacter coli isolates of human and animal origin collected in Rio de Janeiro City, 30 C. jejuni and 35 C. coli isolates from animal sources (n = 45) and human patients with gastroenteritis (n = 20) were genotyped by PCR-based techniques, namely random amplified polymorphic DNA (RAPD-PCR) and enterobacterial repetitive intergenic consensus sequence (ERIC-PCR). RAPD-PCR identified 50 types and ERIC-PCR identified 22 genotypes, among the 65 Campylobacter isolates. Both PCR methods discriminated the C. jejuni and C. coli groups of isolates. Combining the results of both methods, no single genotype was shared between isolates from human and animal sources. Two groups of two C. coli isolates each with identical genotypes were found among poultry and pig isolates. A high level of genetic diversity observed among the Campylobacter isolates suggests lack of overlap between isolates from different sources.     

Occurrence of ɛ-proteobacterial species in rabbits (Oryctolagus cuniculus) reared in intensive and rural farms

In order to investigate the occurrence of Campylobacter, Helicobacter and Arcobacter species in caecal contents of rabbits reared in intensive and rural farms, a total of 87 samples from animals belonging to 29 farms were analysed by both cultural and PCR analyses.