抗生素和肠道细菌对小菜蛾适合度的影响

【目的】小菜蛾是危害十字花科蔬菜的世界性重要害虫。通过分析抗生素对小菜蛾的毒性效应,了解肠道细菌对小菜蛾适合度的影响,有助于更好地阐明小菜蛾肠道细菌的功能。【方法】利用抗生素处理含有高丰度肠道细菌的萝卜苗饲养品系(FZss)小菜蛾幼虫,同时利用抗生素处理饲料饲养的无肠道细菌(SLss)小菜蛾幼虫,分析抗生素及肠道细菌对小菜蛾适合度的影响。【结果】抗生素处理FZss品系小菜蛾导致了小菜蛾发育历期延长,虫重、蛹重、化蛹率、产卵量和成虫寿命降低。利用抗生素处理无肠道菌的SLss品系小菜蛾幼虫,小菜蛾化蛹率和单雌产卵量均显著降低,而对发育历期、虫重和蛹重则无影响。【结论】综合两个研究的结果发现抗生素处理后宿主适合度的降低一方面是由于抗生素的毒性效应导致,另一方面是由于小菜蛾肠道细菌的缺失引起。抗生素的毒性效应主要表现为对化蛹率和单雌产卵量的影响,而肠道细菌则对小菜蛾的发育历期、虫重、蛹重以及成虫寿命具有重要的促进作用。     

Bacteria abundance and diversity of different life stages of Plutella xylostella (Lepidoptera: Plutellidae), revealed by bacteria culture‐dependent and PCR‐DGGE methods

Microbial abundance and diversity of different life stages (fourth instar larvae, pupae and adults) of the diamondback moth, Plutella xylostella L., collected from field and reared in laboratory, were investigated using bacteria culture‐dependent method and PCR‐DGGE analysis based on the sequence of bacteria 16S rRNA V3 region gene. A large quantity of bacteria was found in all life stages of P. xylostella. Field population had higher quantity of bacteria than laboratory population, and larval gut had higher quantity than pupae and adults. Culturable bacteria differed in different life stages of P. xylostella. Twenty‐five different bacterial strains were identified in total, among them 20 strains were presented in larval gut, only 8 strains in pupae and 14 strains in adults were detected. Firmicutes bacteria, Bacillus sp., were the most dominant species in every life stage. 15 distinct bands were obtained from DGGE electrophoresis gel. The sequences blasted in GenBank database showed these bacteria belonged to six different genera. Phylogenetic analysis showed the sequences of the bacteria belonged to the Actinobacteri, Proteobacteria and Firmicutes. Serratia sp. in Proteobacteria was the most abundant species in larval gut. In pupae, unculturable bacteria were the most dominant species, and unculturable bacteria and Serratia sp. were the most dominant species in adults. Our study suggested that a combination of molecular and traditional culturing methods can be effectively used to analyze and to determine the diversity of gut microflora. These known bacteria may play important roles in development of P. xylostella.     

A mid-gut microbiota is not required for the pathogenicity of Bacillus thuringiensis to diamondback moth larvae

The mode of action of the entomopathogenic bacterium Bacillus thuringiensis ( Bt ) remains a matter of debate. Recent reports have claimed that aseptic lepidopteran hosts were not susceptible to Bt and that inoculation with mid-gut bacteria restores pathogenicity. These claims are controversial because larvae were rendered aseptic by consuming antibiotics, although the effect of these antibiotics on Bt was not examined. We tested the generality of the mid-gut bacteria hypothesis in the diamondback moth, Plutella xylostella using properly controlled experiments that investigated the effect of antibiotic consumption and absence of gut microbiota separately. We found that purified Bt toxin and spore/toxin mixtures were fully pathogenic to larvae reared aseptically. Persistence of antibiotics in larval tissues was implicated in reducing host mortality because larval consumption of the antibiotic rifampicin reduced the pathogenicity of rifampicin-sensitive Bt strains but not rifampicin-resistant strains. Inoculating larvae with Enterobacter sp. Mn2 reduced the mortality of larvae feeding on Bt HD-1 and the presence of a culturable gut microbiota also reduced the pathogenicity of the Bt toxin Cry1Ac, in agreement with other studies indicating that an intestinal microbiota can protect taxonomically diverse hosts from pathogen attack. As ingestion of antibiotics suppresses host mortality the vegetative growth of Bt in the host must be important for its pathogenicity. Furthermore, claims that aseptic larvae are not susceptible to Bt must be supported by experiments that control for the effect of administering antibiotics.     

Autochthonous bacterial flora indicated by PCR-DGGE of 16S rRNA gene fragments from the alimentary tract of Costelytra zealandica (Coleoptera: Scarabaeidae)

Aims: To locate and identify putative autochthonous bacteria within the grass grub gut that may have a role in symbiosis. Methods and Results: Polymerase chain reaction-denaturing gradient gel electrophoresis (DGGE) fingerprinting was used to investigate bacterial diversity in the grass grub larval gut. The microbial community profiles from five geographically distinct populations were compared and the influence of feeding was analysed. Bacterial community in the midgut was highly variable between locations and was affected by feeding. The hindgut contained a more diverse but stable bacterial community that was less affected by external conditions. Forty-seven distinct DGGE bands, representing different bacterial genotypes, could be distinguished from all samples, with 34 different bands occurring in the hindgut. The 22 most common bands were isolated and DNA was sequenced. Sequence analysis revealed that most bacteria (16/22) were affiliated to the Clostridiales with the predominant bacteria affiliated to the genus Clostridium. The remaining bacteria were aligned to the Betaproteobacteria, Deltaproteobacteria and Bacteroidetes. Conclusions: The grass grub larva has an autochthonous microflora with predominance of Clostridium spp. in the hindgut. Significance and Impact of the Study: Occurrence of an autocthonous microflora in the grass grub hindgut suggests a symbiotic relationship which could help explain the ability of larval scarabs to feed on recalcitrant organic matter.     

Gut microbiota of the pine weevil degrades conifer diterpenes and increases insect fitness

The pine weevil (Hylobius abietis), a major pest of conifer forests throughout Europe, feeds on the bark and cambium, tissues rich in terpenoid resins that are toxic to many insect herbivores. Here, we report the ability of the pine weevil gut microbiota to degrade the diterpene acids of Norway spruce. The diterpene acid levels present in ingested bark were substantially reduced on passage through the pine weevil gut. This reduction was significantly less upon antibiotic treatment, and supplementing the diet with gut suspensions from untreated insects restored the ability to degrade diterpenes. In addition, cultured bacteria isolated from pine weevil guts were shown to degrade a Norway spruce diterpene acid. In a metagenomic survey of the insect's bacterial community, we were able to annotate several genes of a previously described diterpene degradation (dit) gene cluster. Antibiotic treatment disrupted the core bacterial community of H. abietis guts and eliminated nearly all dit genes concordant with its reduction in diterpene degradation. Pine weevils reared on an artificial diet spiked with diterpenes, but without antibiotics, were found to lay more eggs with a higher hatching rate than weevils raised on diets with antibiotics or without diterpenes. These results suggest that gut symbionts contribute towards host fitness, but not by detoxification of diterpenes, as these compounds do not show toxic effects with or without antibiotics. Rather the ability to thrive in a terpene‐rich environment appears to allow gut microbes to benefit the weevil in other ways, such as increasing the nutritional properties of their diet.     

Delayed larval development in Anopheles mosquitoes deprived of Asaiabacterial symbionts

Background

In recent years, acetic acid bacteria have been shown to be frequently associated with insects, but knowledge on their biological role in the arthropod host is limited. The discovery that acetic acid bacteria of the genus Asaia are a main component of the microbiota of Anopheles stephensi makes this mosquito a useful model for studies on this novel group of symbionts. Here we present experimental results that provide a first evidence for a beneficial role of Asaia in An. stephensi.

Results

Larvae of An. stephensi at different stages were treated with rifampicin, an antibiotic effective on wild-type Asaia spp., and the effects on the larval development were evaluated. Larvae treated with the antibiotic showed a delay in the development and an asynchrony in the appearance of later instars. In larvae treated with rifampicin, but supplemented with a rifampicin-resistant mutant strain of Asaia, larval development was comparable to that of control larvae not exposed to the antibiotic. Analysis of the bacterial diversity of the three mosquito populations confirmed that the level of Asaia was strongly decreased in the antibiotic-treated larvae, since the symbiont was not detectable by PCR-DGGE (denaturing gradient gel electrophoresis), while Asaia was consistently found in insects supplemented with rifampicin plus the antibiotic-resistant mutant in the diet, and in those not exposed to the antibiotic.

Conclusions

The results here reported indicate that Asaia symbionts play a beneficial role in the normal development of An. stephensi larvae.

     

Contribution of gut bacteria to digestion and development of the velvetbean caterpillar, Anticarsia gemmatalis

Bacteria colonies from gut homogenates of fifth instar velvetbean caterpillars (Lepidoptera: Noctuidae) were subjected to antibiotic sensitivity experiments using discs containing 22 antibiotics. The antibiotic tetracycline provided the best results, followed by chloramphenicol. Tetracycline also provided higher inhibition of colony forming units than chloramphenicol and was therefore provided to the caterpillars in increasing diet concentrations to assess the contribution of gut bacteria to their digestion and development. The activity of proteases (general), serine-proteinases and lipases were significantly suppressed by tetracycline. Concentration-inhibition curves were successfully established for tetracycline and this antibiotic was effective in suppressing them, particularly serine-proteinases, suggesting that gut bacteria may significantly contribute with lipid- and mainly protein-digestion in velvetbean caterpillars. Increased diet concentrations of tetracycline led only to mild increase in insect mortality (ca. 20%), with the surviving insects showing faster development (≤4 days) and higher pupa weight (<0.04 mg) with increased concentrations of tetracycline. Therefore, the gut bacteria inhibited by tetracycline does not seem to play a crucial role in the survival and development of the velvetbean caterpillar, but may be important in the adaptation of this pest species to hosts rich in protease inhibitors, such as soybean.     

取食不同食物对小菜蛾幼虫肠道细菌多样性的影响

【目的】植食性昆虫肠道细菌的组成与其食物密切相关。本研究旨在探究小菜蛾Plutella xylostella幼虫肠道细菌多样性与其取食食物之间的关系以及它们之间相互适应的过程。【方法】本研究选取小菜蛾人工饲料品系(S)及其转寄主到结球甘蓝Brassica oleracea var. capitata、结球白菜Brassica rapa subsp. pekinensis和花椰菜Brassica olerocea var. botrytis饲养后第1代(分别为G1C, G1CC和G1WC)和第3代(分别为G3C, G3CC和G3WC)的4龄幼虫,提取小菜蛾肠道细菌基因组DNA,利用Illumina MiSeq二代高通量测序技术,分析其肠道细菌多样性和丰度。【结果】α多样性指数分析发现,取食不同食物的小菜蛾4龄幼虫肠道细菌多样性高低顺序为G1WC>G1CC>S>G1C。在菌群组成上,以人工饲料为食的S样品肠道细菌主要由厚壁菌门(Firmicutes)组成,转寄主植物后的G1C, G1CC和G1WC肠道中厚壁菌门(Firmicutes)相对丰度显著下降,G1C和G1CC小菜蛾肠道中变形菌门(Proteobacteria)相对丰度显著上升成为优势菌群,G1WC肠道中拟杆菌门(Bacteroidetes)成为优势菌群。在寄主植物上连续饲养3代后,与第1代相比,小菜蛾肠道细菌α多样性指数没有显著性改变,但在结球甘蓝和结球白菜上小菜蛾肠道菌群结构却发生了变化,相比G1C,G3C肠道中芽孢杆菌目(Bacillales)的相对丰度显著下降;相比G1CC, G3CC肠道中放线菌门(Proteobacteria)、芽单胞菌门(Gemmatimonadetes)和硝化螺旋菌门(Nitrospirae)的相对丰度均显著上升。【结论】取食人工饲料和不同寄主植物的小菜蛾幼虫肠道细菌多样性和群落构成存在显著差异,寄主植物对小菜蛾肠道微生物的结构组成具有重要的影响,且小菜蛾肠道微生物对寄主植物可能存在一个长期适应的过程。本研究为进一步探讨影响小菜蛾肠道细菌变化的因素,以及后续研究肠道细菌与寄主植物之间的互作奠定了良好的基础。     

基于幼虫粪便的小菜蛾肠道细菌分离鉴定及其抗生素敏感性分析

【目的】昆虫肠道微生物对于其食物消化、生长发育以及环境适应性等方面都具有重要作用,本研究旨在探究小菜蛾 Plutella xylostella (L.)幼虫肠道可培养细菌的菌群结构及抗生素敏感性。【方法】对小菜蛾3龄幼虫粪便进行了细菌分离培养和16S rDNA分子鉴定,并采用纸片扩散法对之进行了药敏试验。【结果】小菜蛾3龄幼虫肠道有5属6种细菌,即蒙氏肠球菌 Enterococcus mundtii、欧文氏菌属 Erwinia 2种、成团泛菌 Pantoea agglomerans 、类芽孢杆菌Paenibacillus sp.和栖稻假单胞菌 Pseudomonas oryzihabitans,均与小菜蛾中肠细菌已知种不同,其中蒙氏肠球菌数量最多;蒙氏肠球菌、桃色欧文氏菌和成团泛菌对青霉素、氨苄青霉素、麦迪霉素、克拉霉素和洁霉素均不敏感,而对磷霉素、万古霉素和强力霉素等表现出相似的敏感性。【结论】小菜蛾幼虫肠道细菌菌群组成具有多样性,且存在天然耐药性现象。本研究为进一步开展小菜蛾幼虫肠道细菌微生物区系及功能细菌的研究提供了菌株材料和研究基础。     

Detection and eradication of endophytic bacteria from micropropagated mint plants

Summary Liquid medium and an enriched agar were used to detect endophytic bacteria in micropropagated mints (Mentha spp.) within 2 to 6 d of inoculation. Bacteria isolated from the cultures were screened on several antibiotic concentrations to determine bactericidal doses. Antibiotics were also tested for phytotoxic effects. Shoot tips from infected plants were treated by immersion in liquid MS medium containing antibiotics either singly or in combination. Streptomycin applied at 1000&#x00B5;g/ml for a period of 10 d was effective and less phytotoxic in a larger number of cases than gentamicin (50&#x00B5;g/ml), neomycin (500&#x00B5;g/ml), or rifampicin (30&#x00B5;g/ml). Mint cultures that tested negative for bacteria after antibiotic treatment were multiplied, retested, and cold-stored for 1 yr or longer. Upon regrowth after storage, 25 of 30 treated cultures (83%) tested negative for bacteria. Of the 25, 8 were successfully treated with streptomycin, 1 with gentamicin, 2 with neomycin, 1 with rifampicin, and 1 with streptomycin and gentamicin; 12 required more than one treatment. An early detection system, initial trial treatment with streptomycin for infected plants, and monitoring of treated cultures successfully reduced the spread of bacterial contamination. Antibiotic treatment in liquid MS medium at pH 6.9 resulted in enhanced bactericidal activity over that seen at pH 5.5.     

4种抗生素对蚯蚓-污泥系统细菌群落结构的影响

蚯蚓可摄食污泥中的有机物,其肠道微生物群落在其分解过程中起着主要的作用。利用赤子爱胜蚓(Eisenia foetida)和人工湿地基质构建蚯蚓-污泥系统,添加氯霉素、四环素、链霉素和青霉素4种抗生素,研究不同抗生素对污泥和蚓粪的细菌群落结构的影响。采用高通量测序技术比较分析污泥和蚓粪的细菌多样性及群落结构变化。结果表明,外加抗生素能够导致污泥的Chao1和ACE指数降低,同时降低拟杆菌门和变形菌门的相对丰度,加入氯霉素和青霉素会增加厚壁菌门的相对丰度,降低酸杆菌门、放线菌门和绿弯菌门的相对丰度,加入四环素和链霉素则与之相反。蚓粪样品中,添加氯霉素和链霉素导致Chao1和ACE指数降低,而添加四环素和青霉素则导致Chao1和ACE指数升高,外加抗生素可降低拟杆菌门的相对丰度,增加放线菌门、变形菌门和疣微菌门的相对丰度。主成分分析(PCA)和聚类分析表明,氯霉素和青霉素对污泥细菌群落影响作用相似,四环素与链霉素效果类似;氯霉素对蚓粪群落结构的影响小于其他抗生素。研究结果显示,抗生素可影响污泥和蚓粪的细菌多样性及群落结构,不同抗生素对污泥和蚓粪的影响程度存在差异。     

The effect of different oral antibiotics on the gastrointestinal microflora of a wild rodent (Aethomys namaquensis)

Gut sterilization via the oral administration of antibiotics facilitates physiological studies of the nutritionally important relationship between intestinal microflora and the host. However, the composition of gut flora is extremely variable, and as a result, the efficacy of antibiotics in achieving gut sterilization varies considerably between species. We tested the effectiveness of three antibiotic cocktails in sterilizing the gut of a rodent pollinator, the Namaqua rock mouse (Aethomys namaquensis). The cocktails were (1) streptomycin sulfate and bacitracin (previously used with domestic mice and rats), (2) chloramphenicol and bacitracin (based on antibiotic screening tests performed on faecal flora) and (3) Baytril 10% oral solution (a veterinary antimicrobial agent containing enrofloxacin). We tested for antibiotic inactivation by determining bacterial viability through fluorescence staining of faecal samples. We also tested techniques to maintain sterility during antibiotic treatment without the benefit of a laminar flow cabinet. Antibiotics were administered orally in food and water consumed ad libitum over 4 consecutive days. Antibiotic effectiveness was assessed by culturing anaerobic bacteria from faecal samples collected before and after each antibiotic treatment. Treatment with Baytril 10% oral solution eliminated or significantly reduced faecal flora, whereas other antibiotics did not. This study clearly demonstrates the importance of testing the effectiveness of antibiotics before their use in studies that involve antibiotic-treated subjects, particularly if these are species previously untested.     

Ultrasonic enhancement of antibiotic action on several species of bacteria

The effect of the antibiotics gentamicin, streptomycin, kanamycin, tetracycline, and ampicillin on planktonic cultures of Enterobacter aerogenes, Serratia marcescens, Salmonella derby, Streptococcus mitis, and Staphylococcus epidermidis with and without an application of 70 kHz ultrasound was studied. The ultrasound was applied at levels that had no inhibitory effect on planktonic cultures of bacteria. Measurements of viability at, above, and below the minimum inhibitory concentration of the above antibiotics on the planktonic cultures of these bacteria showed that a simultaneous application of 70 kHz ultrasound and antibiotic significantly increased the effectiveness of selected antibiotics. Bacterial viability was reduced several orders of magnitude when harmless levels of ultrasound were combined with some antibiotics, especially the aminoglycosides. Similar synergistic effects of combined ultrasound and antibiotic treatment were seen in both Gram-positive and Gram-negative bacteria with several classes of antibiotics. These results may have application in the treatment of bacterial infections normally resistant to some antibiotics.     

Effect of diamide administration on longevity, oxygen consumption, superoxide dismutase, catalase, inorganic peroxides and glutathione in the adult housefly, Musca domestica

The effects of various levels of tetracycline hydrochloride on the feeding habits, larval growth, and the metabolism of lipids, carbohydrates and proteins of Heliothis virescens were studied. Regression analysis showed that larval weight, fatty acids and glycogen decreased exponentially with increasing concentration of antibiotic, whereas protein showed a linear decrease. Larval feeding was initially decreased when antibiotics were added to the diet but there was no difference after 24 hr.     

Intracellular production of Brucella L forms . II. Induction and survival of Brucella abortus L forms in tissue culture

Hatten, Betty A. (The University of Texas Southwestern Medical School, Dallas), and S. Edward Sulkin. Intracellular production of Brucella L forms. II. Induction and survival of Brucella abortus L forms in tissue culture. J. Bacteriol. 91:14-20. 1966.-Intracellular survival of altered brucellae, possibly L forms, was not greatly affected by penicillin or streptomycin in concentrations ranging from 5.0 to 40 mug/ml, but a combination of these two antibiotics (2.5 to 20 mug/ml each) reduced the number of positive L-form cultures. Tetracycline (2.0 mug/ml) decreased the number of positive L-form cultures at about the same rate as combinations of the higher concentrations of penicillin and streptomycin. Various concentrations of tetracycline (0.1 to 2.0 mug/ml) with 5.0 mug/ml of penicillin or streptomycin significantly reduced the number of positive L-form cultures. L forms were recovered for several days after elimination of bacteria from the cultures by all of the antibiotics tested. L-form production was not dependent upon the presence of antibiotics in the culture medium, but they were recovered in greater numbers when bacteria were still present in the hamster kidney cells. Addition of thallium acetate to infected cells (at varying intervals of time after infection) to control bacterial growth and conversion to the L phase during cellular disintegration decreased the number of positive L-form cultures obtained over a 10-day period. Comparison of the antibiotic sensitivity of bacteria recovered from infected tissue culture cells with the stock strain of Brucella abortus indicated that some resistance to penicillin and tetracycline had developed. A marked resistance to streptomycin was observed in those bacteria recovered from cells maintained in the presence of this antibiotic.     

Bacteriophage versus antibiotic therapy on gut bacterial communities of juvenile green turtle,Chelonia mydas

Green turtles are endangered marine herbivorous hindgut fermenters that contribute to a variety of marine ecosystems. Debilitated turtles are often rehabilitated in turtle hospitals. Since accurate diagnosis of disease is difficult, broad-spectrum antibiotics are routinely used as a general treatment, potentially causing collateral damage to the gut microbiome of the patient. Here, we evaluated the concept of the application of bacteriophage (phages) to eliminate targeted intestinal bacteria as an alternative to a broad-spectrum antibiotic (enrofloxacin) in clinically healthy, captive green turtles. Additionally, the impact of a broad-spectrum antibiotic (enrofloxacin) and phage therapy on the gut bacterial communities of green turtles was evaluated. Gut bacterial communities in faecal samples were analysed by sequencing the V1-V3 regions of the bacterial 16S rRNA. Bacteria-specific phage cocktails significantly (P < 0.05) reduced targeted Acinetobacter in phage-treated turtles during the therapy. Compared to control, no significant difference was observed in the bacterial diversity and compositions in phage-treated turtles. In contrast, bacterial diversity was significantly (P < 0.05) reduced in antibiotic-treated turtles at day 15 and throughout the trial. The alteration in the bacterial microbiota of antibiotic-treated turtles was largely due to an increase in abundance of Gram-positive Firmicutes and a concurrent decrease in Gram-negative Bacteroidetes, Proteobacteria and Verrucomicrobia. Additionally, we observed the relative abundance of several bacteria at lower taxonomic level was much less affected by phages than by antibiotics. These data offer the proof of concept of phage therapy to manipulate transient as well as indigenous bacterial flora in gut-related dysbiosis of turtles.     

A mutant of Bacillus thuringiensis var. israelensis (B.t.i.) resistant to antibiotics

Summary PST, a spontaneous mutant of Bacillus thuringiensis var. israelensis (B.t.i.) resistant to penicillin, streptomycin and tetracycline was isolated by serial selections. In the absence of antibiotics it showed genetic stability for 16 generations. Mosquito larvicidal activity of PST was similar to that of B.t.i., its parental strain. It also maintained the specific antigenicity of B.t.i. although its rate of growth was somewhat lower, a generation time of 55 min for PST vs. 38 min for B.t.i. Cell concentration plays a major role in the phenomenon of PST resistance to penicillin.This antibiotic resistant mutant of b.t.i. provides us with an efficient tool to trace B.t.i. among the indigenous bacteria present in septic habitats in the field as well as inside the larval gut.     

Spatial and temporal variability of antibiotic resistance in freshwater bacterial assemblages

Abstract Spatial and temporal variability in antibiotic resistance was examined in bacterial assemblages from streams and ponds on the US Department of Energy's Savannah River Site (SRS) in South Carolina. Sites sampled have been impacted to varying degrees by contamination with organic compounds, heavy metals, and radioactive materials because of production of nuclear materials on the site. Antibiotic resistance in the culturable portion of the bacterial assemblage was determined from coloby formation on media containing antibiotics. Eight antibiotics, chloramphenicol, cycloserine, kanamycin, neomycin, novobiocin, rifampicin, streptomycin, and tetracycline, were used at concentrations of 50 and 200 μg ml⁻¹. Statistically significant differences in frequency of antibiotic resistance were observed among sites and among dates at a single site. Bacterial densities (total and culturable), dissolved organic carbon (DOC) concentration, and human impact also varied among sites but bore no overall relationship to resistance frequency. SRS operations did not have a detectable impact on antibiotic resistance.     

Development of bacterial and bifidobacterial communities in feces of newborn babies

Microbial 16S rDNA from babies' fecal samples were amplified by PCR, and analysed by denaturing gradient gel electrophoresis (DGGE), cloning and sequencing. PCR-DGGE profiles were used to follow in time the colonization of the intestine by bacteria. Four healthy babies, one baby who received antibiotics and their parents participated to the present study to determine the extent to which administration of antibiotics can modify the bacterial colonization of neonatal human gut and verify the influence of parental factors on the formation of the fecal bacterial community. In the healthy babies, Escherichia coli or bacteria belonging to Clostridium spp. were the initial colonizers rapidly followed by Bifidobacterium, Bacteroides, Clostridium, Streptococcus, Enterococcus and Actinomyces. Bifidobacterium species appeared already after five days in the breast-fed babies while there was a delay in the baby who received a formula based diet during only one day after birth. In each baby two or three bifidobacterial species including B. infantis were found. The observed variations in species were not associated with the feeding changes. The comparison of DGGE profiles of the babies and their parents patterns showed bands with equal migration suggesting a vertical transmission determined by genetic and environmental factors. The brief appearance of pioneer bacteria determined as being E. coli and Enterococcus spp. in the profile from the baby under antibiotic therapy, was succeeded by a small stable community consisting of Ruminococcus species. No Bifidobacterium sequences were detectable in this antibiotic-treated baby in spite of a partly breast-milk diet.     

Susceptibility to Antibiotics of Vibrio sp. AO1 Growing in Pure Culture or in Association with its Hydroid Host Aglaophenia octodonta (Cnidaria, Hydrozoa)

Vibrio harveyi is the major causal organism of vibriosis, causing potential devastation to diverse ranges of marine invertebrates over a wide geographical area. These microorganisms, however, are phenotypically diverse, and many of the isolates are also resistant to multiple antibiotics. In a previous study, we described a previously unknown association between Vibrio sp. AO1, a luminous bacterium related to the species V. harveyi, and the benthic hydrozoan Aglaophenia octodonta. In this study, we analyzed the susceptibility to antibiotics (ampicillin, streptomycin, tetracycline, or co-trimoxazole = mix of sulfamethoxazole and trimetoprim) of Vibrio sp. AO1 growing in pure culture or in association with its hydroid host by using microcosm experiments. The results of minimum inhibitory concentration (MIC) experiments demonstrated that Vibrio sp. AO1 was highly resistant to ampicillin and streptomycin in pure culture. Nevertheless, these antibiotics, when used at sub-MIC values, significantly reduced the hydroid fluorescence. Co-trimoxazole showed the highest inhibitory effect on fluorescence of A. octodonta. However, in all treatments, the fluorescence was reduced after 48 h, but never disappeared completely around the folds along the hydrocaulus and at the base of the hydrothecae of A. octodonta when the antibiotic was used at concentration completely inhibiting growth in vitro. The apparent discrepancy between the MIC data and the fluorescence patterns may be due to either heterogeneity of the bacterial population in terms of antibiotic susceptibility or specific chemical–physical conditions of the hydroid microenvironment that may decrease the antibiotic susceptibility of the whole population. The latter hypothesis is supported by scanning electron microscope evidence for development of bacterial biofilm on the hydroid surface. On the basis of the results obtained, we infer that A. octodonta might behave as a reservoir of antibiotic multiresistant bacteria, increasing the risk of their transfer into aquaculture farms.