Effects of Leishmania major infection on the gut microbiome of resistant and susceptible mice.

Cutaneous leishmaniasis, a parasitic disease caused by Leishmania major, is a widely frequent form in humans. To explore the importance of the host gut microbiota and to investigate its changes during L. major infection, two different groups of mouse models were assessed. The microbiome of two parts of the host gut-ileum and colon-from infected and non-infected mice were characterised by sequencing of 16S rDNA using an Ion Torrent PGM platform. Microbiome analysis was performed to reveal changes related to the susceptibility and the genetics of mice strains in two different gut compartments and to compare the results between infected and non-infected mice. The results showed that Leishmania infection affects mainly the ileum microbiota, whereas the colon bacterial community was more stable. Different biomarkers were determined in the gut microbiota of infected resistant mice and infected susceptible mice using LEfSe analysis. Lactobacillaceae was associated with resistance in the colon microbiota of all resistant mice strains infected with L. major. Genes related to xenobiotic biodegradation and metabolism and amino acid metabolism were primarily enriched in the small intestine microbiome of resistant strains, while genes associated with carbohydrate metabolism and glycan biosynthesis and metabolism were most abundant in the gut microbiome of the infected susceptible mice. These results should improve our understanding of host-parasite interaction and provide important insights into the effect of leishmaniasis on the gut microbiota. Also, this study highlights the role of host genetic variation in shaping the diversity and composition of the gut microbiome. KEY POINTS: • Leishmaniasis may affect mainly the ileum microbiota while colon microbiota was more stable. • Biomarkers related with resistance or susceptibility were determined in the gut microbiota of mice. • Several pathways were predicted to be upregulated in the gut microbiota of resistant or susceptible mice.

Dysbiosis of skin microbiome and gut microbiome in melanoma progression.

BACKGROUND: The microbiome alterations are associated with cancer growth and may influence the immune system and response to therapy. Particularly, the gut microbiome has been recently shown to modulate response to melanoma immunotherapy. However, the role of the skin microbiome has not been well explored in the skin tumour microenvironment and the link between the gut microbiome and skin microbiome has not been investigated in melanoma progression. Therefore, the aim of the present study was to examine associations between dysbiosis in the skin and gut microbiome and the melanoma growth using MeLiM porcine model of melanoma progression and spontaneous regression. RESULTS: Parallel analysis of cutaneous microbiota and faecal microbiota of the same individuals was performed in 8 to 12 weeks old MeLiM piglets. The bacterial composition of samples was analysed by high throughput sequencing of the V4-V5 region of the 16S rRNA gene. A significant difference in microbiome diversity and richness between melanoma tissue and healthy skin and between the faecal microbiome of MeLiM piglets and control piglets were observed. Both Principal Coordinate Analysis and Non-metric multidimensional scaling revealed dissimilarities between different bacterial communities. Linear discriminant analysis effect size at the genus level determined different potential biomarkers in multiple bacterial communities. Lactobacillus, Clostridium sensu stricto 1 and Corynebacterium 1 were the most discriminately higher genera in the healthy skin microbiome, while Fusobacterium, Trueperella, Staphylococcus, Streptococcus and Bacteroides were discriminately abundant in melanoma tissue microbiome. Bacteroides, Fusobacterium and Escherichia-Shigella were associated with the faecal microbiota of MeLiM piglets. Potential functional pathways analysis based on the KEGG database indicated significant differences in the predicted profile metabolisms between the healthy skin microbiome and melanoma tissue microbiome. The faecal microbiome of MeLiM piglets was enriched by genes related to membrane transports pathways allowing for the increase of intestinal permeability and alteration of the intestinal mucosal barrier. CONCLUSION: The associations between melanoma progression and dysbiosis in the skin microbiome as well as dysbiosis in the gut microbiome were identified. Results provide promising information for further studies on the local skin and gut microbiome involvement in melanoma progression and may support the development of new therapeutic approaches.

Bifidobacterium canis sp. nov., a novel member of the Bifidobacterium pseudolongum phylogenetic group isolated from faeces of a dog (Canis lupus f. familiaris).

A fructose-6-phosphate phosphoketolase-positive strain (GSD1FST) was isolated from a faecal sample of a 3 weeks old German Shepherd dog. The closest related taxa to isolate GSD1FST based on results from the EZBioCloud database were Bifidobacterium animalis subsp. animalis ATCC 25527T, Bifidobacterium animalis subsp. lactis DSM 10140T and Bifidobacterium anseris LMG 30189T, belonging to the Bifidobacterium pseudolongum phylogenetic group. The resulting 16S rRNA gene identities (compared length of 1454 nucleotides) towards these taxa were 97.30, 97.23 and 97.09 %, respectively. The pairwise similarities of strain GSD1FST using argS, atpA, fusA, hsp60, pyrG, rpsC, thrS and xfp gene fragments to all valid representatives of the B. pseudolongum phylogenetic group were in the concatenated range of 83.08-88.34 %. Phylogenomic analysis based on whole-genome methods such as average nucleotide identity revealed that bifidobacterial strain GSD1FST exhibits close phylogenetic relatedness (88.17 %) to Bifidobacetrium cuniculi LMG 10738T. Genotypic characteristics and phylogenetic analyses based on nine molecular markers, as well as genomic and comparative phenotypic analyses, clearly proved that the evaluated strain should be considered as representing a novel species within the B. pseudolongum phylogenetic group named as Bifidobacterium canis sp. nov. (GSD1FST=DSM 105923T=LMG 30345T=CCM 8806T).

[Two case reports of chronic hepatitis C retreatment].

In a group of 211 patients with chronic hepatitis C treated with direct-acting antivirals, four experienced therapy failure. Two patients, one originally treated with dasabuvir/ombitasvir/paritaprevir/ritonavir and the other with glecaprevir/pibrentasvir, received a triple combination of sofosbuvir, velpatasvir and voxilaprevir for 12 weeks. Following the retreatment, both patients were permanently virus-free.

Molecular Epidemiology of Hantaviruses in the Czech Republic.

During 2008-2018, we collected samples from rodents and patients throughout the Czech Republic and characterized hantavirus isolates. We detected Dobrava-Belgrade and Puumala orthohantaviruses in patients and Dobrava-Belgrade, Tula, and Seewis orthohantaviruses in rodents. Increased knowledge of eco-epidemiology of hantaviruses will improve awareness among physicians and better outcomes of patients.

Comparison of enzymatic activities and proteomic profiles of Butyrivibrio fibrisolvens grown on different carbon sources.

BACKGROUND: The rumen microbiota is one of the most complex consortia of anaerobes, involving archaea, bacteria, protozoa, fungi and phages. They are very effective at utilizing plant polysaccharides, especially cellulose and hemicelluloses. The most important hemicellulose decomposers are clustered with the genus Butyrivibrio. As the related species differ in their range of hydrolytic activities and substrate preferences, Butyrivibrio fibrisolvens was selected as one of the most effective isolates and thus suitable for proteomic studies on substrate comparisons in the extracellular fraction. The B. fibrisolvens genome is the biggest in the butyrivibria cluster and is focused on "environmental information processing" and "carbohydrate metabolism". METHODS: The study of the effect of carbon source on B. fibrisolvens 3071 was based on cultures grown on four substrates: xylose, glucose, xylan, xylan with 25% glucose. The enzymatic activities were studied by spectrophotometric and zymogram methods. Proteomic study was based on genomics, 2D electrophoresis and nLC/MS (Bruker Daltonics) analysis. RESULTS: Extracellular ß-endoxylanase as well as xylan ß-xylosidase activities were induced with xylan. The presence of the xylan polymer induced hemicellulolytic enzymes and increased the protein fraction in the interval from 40 to 80 kDa. 2D electrophoresis with nLC/MS analysis of extracellular B. fibrisolvens 3071 proteins found 14 diverse proteins with significantly different expression on the tested substrates. CONCLUSION: The comparison of four carbon sources resulted in the main significant changes in B. fibrisolvens proteome occurring outside the fibrolytic cluster of proteins. The affected proteins mainly belonged to the glycolysis and protein synthesis cluster.

Effect of Selected Stilbenoids on Human Fecal Microbiota.

Dietary phenolics or polyphenols are mostly metabolized by the human gut microbiota. These metabolites appear to confer the beneficial health effects attributed to phenolics. Microbial composition affects the type of metabolites produced. Reciprocally, phenolics modulate microbial composition. Understanding this relationship could be used to positively impact health by phenolic supplementation and thus create favorable colonic conditions. This study explored the effect of six stilbenoids (batatasin III, oxyresveratrol, piceatannol, pinostilbene, resveratrol, thunalbene) on the gut microbiota composition. Stilbenoids were anaerobically fermented with fecal bacteria from four donors, samples were collected at 0 and 24 h, and effects on the microbiota were assessed by 16S rRNA gene sequencing. Statistical tests identified affected microbes at three taxonomic levels. Observed microbial composition modulation by stilbenoids included a decrease in the Firmicutes to Bacteroidetes ratio, a decrease in the relative abundance of strains from the genus Clostridium, and effects on the family Lachnospiraceae. A frequently observed effect was a further decrease of the relative abundance when compared to the control. An opposite effect to the control was observed for Faecalibacterium prausnitzii, whose relative abundance increased. Observed effects were more frequently attributed to resveratrol and piceatannol, followed by thunalbene and batatasin III.

[Use of PCR for diagnosing cat-scratch disease].

OBJECTIVE: The diagnosis of Bartonella henselae by polymerase chain reaction (PCR) in lymph nodes removed in 10 patients with serologically confirmed evidence cat-scratch disease. MATERIAL AND METHODS: The 2015-2018 group consisted of 10 patients with serologically confirmed cat-scratch disease, all of them having positive IgG antibodies and 6 patients also positive IgM antibodies against B. henselae. The group included 4 men and 6 women, 7 children and 3 adults, aged 5-52 years. Eleven lymph nodes obtained from the 10 patients were formalin-fixed paraffin-embedded. Variants of granulomatous inflammation were found in 9 patients; a 13-year-old boy had Hodgkin's lymphoma. DNA isolation was performed with cobas® DNA Sample Preparation Kit (Roche). DNA of Bartonella spp. was detected by real-time PCR with BactoReal® Kit Bartonella spp. (Ingenetix) detecting the gltA gene specific for the genus Bartonella. RESULTS: Four of the 10 patients tested positive or borderline positive for Bartonella when their histological material was analyzed by PCR. One patient with 2 lymph nodes examined showed a positive result for only 1 lymph node. One adult male had a positive result; three children showed borderline positive results. Of those, two patients had suppurative granulomatous and the other 2 patients had necrotizing suppurative granulomatous inflammation as histological findings. All 4 patients had positive IgM antibodies against B. henselae. The boy with lymphoma had a negative PCR result. CONCLUSION: Serological tests combined with histological examination of lymph nodes and PCR may improve the diagnosis of cat- scratch disease.

[Cryptic Aspergillus species - a case report of chronic cavitary pulmonary Aspergillus lentulus infection in a heart transplant recipient].

Cryptic species within the section Fumigati, that is Aspergillus fumigatus-like species, are increasingly reported in the literature as causative agents of invasive aspergillosis (IA) in both humans and animals. Their detection and proper identification are important, but even more important is to determine the susceptibility profile (minimum inhibitory concentrations, MICs) of the isolate to antifungals using appropriate methods. Cryptic species often demonstrate elevated MICs to drugs recommended for IA therapy such as voriconazole or amphotericin B. Presented is a case of pulmonary aspergillosis in a 63-year-old male heart transplant recipient. Aspergillus lentulus with reduced susceptibility to voriconazole and amphotericin B was identified as the causative agent of the infection using culture and DNA sequencing. Susceptibility to antifungals was confirmed by the standard EUCAST-AFST methods. Based on MIC values obtained in vitro, therapy was switched from voriconazole to posaconazole with excellent clinical effects. To the best of our knowledge, this is the first reported case of A. lentulus infection treated with posaconazole and, moreover, a successful one.

A comparison of methanogens of different regions of the equine hindgut.

The diversity of the methanogenic archaea associated with the six segments of the horse and donkey hindgut (caecum, right ventral colon, left ventral colon, left dorsal colon, right dorsal colon, and rectum) was analyzed using 16S rDNA gene clone library. A total of 641 positive clones, 321 from the horse and 320 from the donkey hindgut, were examined by the RFLP, revealing 9 different ribotypes, 8 in the horse and 5 in the donkey hindgut. In both the animals Methanobacteriales (Methanobrevibacter-like sequences) and Methanomicrobiales (Methanocorpusculum-like sequences) were detected as the dominant orders followed by the uncultured Methanomassiliicoccales. The composition of the equine archaeal community was found to be dependent on the gut region. In both the two animals no Methanobrevibacter-like clones were detected in the caeca, which were instead inhabited by the Methanocorpusculum-like archeons. The Methanosarcinales were found only in distal regions of the horse hindgut.

[Vertical hepatitis C transmission - authors' experiences].

BACKGROUND: Vertical hepatitis C virus (HCV) transmission and persistence of anti-HCV antibodies were retrospectively investigated since 1999 in a group of 244 children whose mothers had a history of hepatitis C. MATERIAL AND METHODS: Initial examinations performed in most children at 6 months of age included the determination of anti-HCV antibodies, HCV nucleic acid (HCV RNA), and anti-HIV antibodies, with all children being negative for HIV. Further examinations with investigation of anti-HCV and HCV RNA were performed at half-year intervals until the disappearance of anti-HCV antibodies. Vertical HCV transmission was defined by HCV RNA positivity in at least 2 venous blood samples or at least two positive anti-HCV results in a child over 3 years of age. RESULTS: Vertical HCV transmission was detected in 11 out of 244 children (4.5%). Only 2 children spontaneously cleared HCV; positive anti-HCV antibodies were last detected when they were 8 years old. Chronic hepatitis C developed in 9 children, four of whom were infected with genotype 1b, 3 children with genotype 3a, one with genotype 1a, and the last one with genotypes 1a and 4. Antiviral treatment including conventional or pegylated interferon, or ribavirin, was administered to 3 children, with sustained elimination of the virus in 2 children. Although the proportion of children with positive anti-HCV antibodies declined gradually, anti-HCV positivity was reported in 6 uninfected children at 18 months of age but in none of them at the age of 2 years. CONCLUSIONS: Vertical transmission of HCV was found in 11 out of 244 children; chronic hepatitis C was detected in 9 children; uninfected children cleared anti-HCV antibodies by 2 years of age.

Reclassification of Eubacterium rectale (Hauduroy et al. 1937) Prévot 1938 in a new genus Agathobacter gen. nov. as Agathobacter rectalis comb. nov., and description of Agathobacter ruminis sp. nov., isolated from the rumen contents of sheep and cows.

Three strains of a butyrate-producing bacterium were isolated from the rumen contents of grazing sheep and cows. The strains were anaerobic, with Gram-positive cell walls, straight-to-slightly-curved, rod-shaped, non-spore-forming and single flagellate. C14 : 1, C14 : 0, C16 : 0 and C16 : 1 were the predominant fatty acids. The cell-wall peptidoglycan type was A1γ. The DNA G+C content varied from 41.4 to 42.2 mol%. 16S rRNA gene sequence similarities between the isolates and Eubacterium rectale, Roseburia hominis and Roseburia intestinalis were found to be 96, 95 and 95 %, respectively. The phylogenetic tree showed that the strains constituted a different taxon, separate from other taxa with validly published names and forming a cluster with strains of Eubacterium rectale. On the basis of phenotypic, chemotaxonomic and phylogenetic results (16S RNA, dnaK, groEL, atpA genes), the isolates are considered to represent a novel species of a new genus of the family Lachnospiraceae, for which the name Agathobacter ruminis gen. nov., sp. nov. is proposed (type strain JK623T = DSM 29029T = LMG 28559T). We also propose the transfer of Eubacterium rectale to the new genus as Agathobacter rectalis gen. nov., comb nov. This new genus represents saccharoclastic, chemo-organotrophic and obligatory anaerobic, non-spore-forming rods with Gram-positive membrane. The main fermentation products on peptone yeast glucose (PYG) medium were butyrate, acetate, hydrogen and lactate. The type species of the genus is Agathobacter rectalis gen. nov., comb nov. (Prévot, 1938) with type strain ATCC 33656T ( = JCM 17463T).

Analysis of the rumen bacterial diversity of goats during shift from forage to concentrate diet.

High-grain feeding used in the animal production is known to affect the host rumen bacterial community, but our understanding of consequent changes in goats is limited. This study was therefore aimed to evaluate bacterial population dynamics during 20 days adaptation of 4 ruminally cannulated goats to the high-grain diet (grain: hay - ratio of 40:60). The dietary transition of goats from the forage to the high-grain-diet resulted in the significant decrease of rumen fluid pH, which was however still higher than value established for acute or subacute ruminal acidosis was not diagnosed in studied animals. DGGE analysis demonstrated distinct ruminal microbial populations in hay-fed and grain-fed animals, but the substantial animal-to-animal variation were detected. Quantitative PCR showed for grain-fed animals significantly higher number of bacteria belonging to Clostridium leptum group at 10 days after the incorporation of corn into the diet and significantly lower concentration of bacteria belonging to Actinobacteria phylum at the day 20 after dietary change. Taxonomic distribution analysed by NGS at day 20 revealed the similar prevalence of the phyla Firmicutes and Bacteroidetes in all goats, significantly higher presence of the unclassified genus of groups of Bacteroidales and Ruminococcaceae in grain-fed animals and significantly higher presence the genus Prevotella and Butyrivibrio in the forage-fed animals. The three different culture-independent methods used in this study show that high proportion of concentrate in goat diet does not induce any serious disturbance of their rumen ecosystem and indicate the good adaptive response of caprine ruminal bacteria to incorporation of corn into the diet.

Gut microbiome composition and metabolomic profiles of wild western lowland gorillas (Gorilla gorilla gorilla) reflect host ecology.

The metabolic activities of gut microbes significantly influence host physiology; thus, characterizing the forces that modulate this micro-ecosystem is key to understanding mammalian biology and fitness. To investigate the gut microbiome of wild primates and determine how these microbial communities respond to the host's external environment, we characterized faecal bacterial communities and, for the first time, gut metabolomes of four wild lowland gorilla groups in the Dzanga-Sangha Protected Areas, Central African Republic. Results show that geographical range may be an important modulator of the gut microbiomes and metabolomes of these gorilla groups. Distinctions seemed to relate to feeding behaviour, implying energy harvest through increased fruit consumption or fermentation of highly fibrous foods. These observations were supported by differential abundance of metabolites and bacterial taxa associated with the metabolism of cellulose, phenolics, organic acids, simple sugars, lipids and sterols between gorillas occupying different geographical ranges. Additionally, the gut microbiomes of a gorilla group under increased anthropogenic pressure could always be distinguished from that of all other groups. By characterizing the interplay between environment, behaviour, diet and symbiotic gut microbes, we present an alternative perspective on primate ecology and on the forces that shape the gut microbiomes of wild primates from an evolutionary context.

Effect of DNA extraction and sample preservation method on rumen bacterial population.

The comparison of the bacterial profile of intracellular (iDNA) and extracellular DNA (eDNA) isolated from cow rumen content stored under different conditions was conducted. The influence of rumen fluid treatment (cheesecloth squeezed, centrifuged, filtered), storage temperature (RT, -80 °C) and cryoprotectants (PBS-glycerol, ethanol) on quality and quantity parameters of extracted DNA was evaluated by bacterial DGGE analysis, real-time PCR quantification and metabarcoding approach using high-throughput sequencing. Samples clustered according to the type of extracted DNA due to considerable differences between iDNA and eDNA bacterial profiles, while storage temperature and cryoprotectants additives had little effect on sample clustering. The numbers of Firmicutes and Bacteroidetes were lower (P < 0.01) in eDNA samples. The qPCR indicated significantly higher amount of Firmicutes in iDNA sample frozen with glycerol (P < 0.01). Deep sequencing analysis of iDNA samples revealed the prevalence of Bacteroidetes and similarity of samples frozen with and without cryoprotectants, which differed from sample stored with ethanol at room temperature. Centrifugation and consequent filtration of rumen fluid subjected to the eDNA isolation procedure considerably changed the ratio of molecular operational taxonomic units (MOTUs) of Bacteroidetes and Firmicutes. Intracellular DNA extraction using bead-beating method from cheesecloth sieved rumen content mixed with PBS-glycerol and stored at -80 °C was found as the optimal method to study ruminal bacterial profile.

Gut microbiota in relationship to diabetes mellitus and its late complications with a focus on diabetic foot syndrome: A review.

Diabetes mellitus is a chronic disease affecting glucose metabolism. The pathophysiological reactions underpinning the disease can lead to the development of late diabetes complications. The gut microbiota plays important roles in weight regulation and the maintenance of a healthy digestive system. Obesity, diabetes mellitus, diabetic retinopathy, diabetic nephropathy and diabetic neuropathy are all associated with a microbial imbalance in the gut. Modern technical equipment and advanced diagnostic procedures, including xmolecular methods, are commonly used to detect both quantitative and qualitative changes in the gut microbiota. This review summarises collective knowledge on the role of the gut microbiota in both types of diabetes mellitus and their late complications, with a particular focus on diabetic foot syndrome.

Microbiome composition and presence of cultivable commensal groups of Southern Tamanduas (Tamandua tetradactyla) varies with captive conditions.

Southern Tamanduas (Tamandua tetradactyla) belong to the specialized placental myrmecophages. There is not much information about their intestinal microbiome. Moreover, due to their food specialization, it is difficult to create an adequate diet under breeding conditions. Therefore, we used 16S rDNA amplicon sequencing to analyze the fecal microbiome of captive Southern Tamanduas from four locations in the Czech Republic and evaluated the impact of the incoming diet and facility conditions on microbiome composition. Together with the microbiome analysis, we also quantified and identified cultivable commensals. The anteater fecal microbiome was dominated by the phyla Bacillota and Bacteroidota, while Pseudomonadota, Spirochaetota, and Actinobacteriota were less abundant. At the taxonomic family level, Lachnospiraceae, Prevotellaceae, Bacteroidaceae, Oscillospiraceae, Erysipelotrichaceae, Spirochaetaceae, Ruminococcaceae, Leuconostocaceae, and Streptococcaceae were mainly represented in the fecal microbiome of animals from all locations. Interestingly, Lactobacillaceae dominated in the location with a zoo-made diet. These animals also had significantly lower diversity of gut microbiome in comparison with animals from other locations fed mainly with a complete commercial diet. Moreover, captive conditions of analyzed anteater included other factors such as the enrichment of the diet with insect-based products, probiotic interventions, the presence of other animals in the exposure, which can potentially affect the composition of the microbiome and cultivable microbes. In total, 63 bacterial species from beneficial commensal to opportunistic pathogen were isolated and identified using MALDI-TOF MS in the set of more than one thousand selected isolates. Half of the detected species were present in the fecal microbiota of most animals, the rest varied across animals and locations.

Defined Pig Microbiota Mixture as Promising Strategy against Salmonellosis in Gnotobiotic Piglets.

Probiotics are a potential strategy for salmonellosis control. A defined pig microbiota (DPM) mixture of nine bacterial strains previously exhibited probiotic and anti-Salmonella properties in vitro. Therefore, we evaluated its gut colonization ability and protection effect against S. typhimurium LT2-induced infection in the gnotobiotic piglet model. The DPM mixture successfully colonized the piglet gut and was stable and safe until the end of the experiment. The colon was inhabited by about 9 log CFU g-1 with a significant representation of bifidobacteria and lactobacilli compared to ileal levels around 7-8 log CFU g-1. Spore-forming clostridia and bacilli seemed to inhabit the environment only temporarily. The bacterial consortium contributed to the colonization of the gut at an entire length. The amplicon profile analysis supported the cultivation trend with a considerable representation of lactobacilli with bacilli in the ileum and bifidobacteria with clostridia in the colon. Although there was no significant Salmonella-positive elimination, it seems that the administered bacteria conferred the protection of infected piglets because of the slowed delayed infection manifestation without translocations of Salmonella cells to the blood circulation. Due to its colonization stability and potential protective anti-Salmonella traits, the DPM mixture has promising potential in pig production applications. However, advanced immunological tests are needed.

Does PAD and microcirculation status impact the tissue availability of intravenously administered antibiotics in patients with infected diabetic foot? Results of the DFIATIM substudy.

Aims/hypothesis: The aim of this substudy (Eudra CT No:2019-001997-27)was to assess ATB availability in patients with infected diabetic foot ulcers(IDFUs)in the context of microcirculation and macrocirculation status. Methods: For this substudy, we enrolled 23 patients with IDFU. Patients were treated with boluses of amoxicillin/clavulanic acid(AMC)(12patients) or ceftazidime(CTZ)(11patients). After induction of a steady ATB state, microdialysis was performed near the IDFU. Tissue fluid samples from the foot and blood samples from peripheral blood were taken within 6 hours. ATB potential efficacy was assessed by evaluating the maximum serum and tissue ATB concentrations(Cmax and Cmax-tissue)and the percentage of time the unbound drug tissue concentration exceeds the minimum inhibitory concentration (MIC)(≥100% tissue and ≥50%/60% tissue fT>MIC). Vascular status was assessed by triplex ultrasound, ankle-brachial and toe-brachial index tests, occlusive plethysmography comprising two arterial flow phases, and transcutaneous oxygen pressure(TcPO2). Results: Following bolus administration, the Cmax of AMC was 91.8 ± 52.5 µgmL-1 and the Cmax-tissue of AMC was 7.25 ± 4.5 µgmL-1(P<0.001). The Cmax for CTZ was 186.8 ± 44.1 µgmL-1 and the Cmax-tissue of CTZ was 18.6 ± 7.4 µgmL-1(P<0.0001). Additionally, 67% of patients treated with AMC and 55% of those treated with CTZ achieved tissue fT>MIC levels exceeding 50% and 60%, respectively. We observed positive correlations between both Cmax-tissue and AUCtissue and arterial flow. Specifically, the correlation coefficient for the first phase was r=0.42; (P=0.045), and for the second phase, it was r=0.55(P=0.01)and r=0.5(P=0.021). Conclusions: Bactericidal activity proved satisfactory in only half to two-thirds of patients with IDFUs, an outcome that appears to correlate primarily with arterial flow.

Tula hantavirus infection in immunocompromised host, Czech Republic.

We report molecular evidence of Tula hantavirus as an etiologic agent of pulmonary-renal syndrome in an immunocompromised patient. Acute hantavirus infection was confirmed by using serologic and molecular methods. Sequencing revealed Tula virus genome RNA in the patient's blood. This case shows that Tula virus can cause serious disease in humans.