Camel Streptococcus agalactiae populations are associated with specific disease complexes and acquired the tetracycline resistance gene tetM via a Tn916-like element

Camels are the most valuable livestock species in the Horn of Africa and play a pivotal role in the nutritional sustainability for millions of people. Their health status is therefore of utmost importance for the people living in this region. Streptococcus agalactiae, a Group B Streptococcus (GBS), is an important camel pathogen. Here we present the first epidemiological study based on genetic and phenotypic data from African camel derived GBS. Ninety-two GBS were characterized using multilocus sequence typing (MLST), capsular polysaccharide typing and in vitro antimicrobial susceptibility testing. We analysed the GBS using Bayesian linkage, phylogenetic and minimum spanning tree analyses and compared them with human GBS from East Africa in order to investigate the level of genetic exchange between GBS populations in the region. Camel GBS sequence types (STs) were distinct from other STs reported so far. We mapped specific STs and capsular types to major disease complexes caused by GBS. Widespread resistance (34%) to tetracycline was associated with acquisition of the tetM gene that is carried on a Tn916-like element, and observed primarily among GBS isolated from mastitis. The presence of tetM within different MLST clades suggests acquisition on multiple occasions. Wound infections and mastitis in camels associated with GBS are widespread and should ideally be treated with antimicrobials other than tetracycline in East Africa.     

Physical carbon‐sequestration mechanisms under special consideration of soil wettability

The protective impact of aggregation on microbial degradation through separation has been described frequently, especially for biotically formed aggregates. However, to date little information exists on the effects of organic‐matter (OM) quantity and OM quality on physical protection, i.e., reduced degradability by microorganisms caused by physical factors. In the present paper, we hypothesize that soil wettability, which is significantly influenced by OM, may act as a key factor for OM stabilization as it controls the microbial accessibility for water, nutrients, and oxygen in three‐phase systems like soil. Based on this hypothesis, the first objective is to evaluate new findings on the organization of organo‐mineral complexes at the nanoscale as one of the processes creating water‐repellent coatings on mineral surfaces. The second objective is to quantify the degree of alteration of coated surfaces with regard to water repellence. We introduce a recently developed trial that combines FTIR spectra with contact‐angle data as the link between chemical composition of OM and the physical wetting behavior of soil particles. In addition to characterizing the wetting properties of OM coatings, we discuss the implications of water‐repellent surfaces for different physical protection mechanisms of OM. For typical minerals, the OM loading on mineral surfaces is patchy, whereas OM forms nanoscaled micro‐aggregates together with metal oxides and hydroxides and with layered clay minerals. Such small aggregates may efficiently stabilize OM against microbial decomposition. However, despite the patchy structure of OM coating, we observed a relation between the chemical composition of OM and wettability. A higher hydrophobicity of the OM appears to stabilize the organic C in soil, either caused by a specific reduced biodegradability of OM or indirectly caused by increased aggregate stability. In partly saturated nonaggregated soil, the specific distribution of the pore water appears to further affect the mineralization of OM as a function of wettability. We conclude that the wettability of OM, quantified by the contact angle, links the chemical structure of OM with a bundle of physical soil properties and that reduced wettability results in the stabilization of OM in soils.     

Improved RP‐HPLC and anion‐exchange chromatography methods for the determination of amino acids and carbohydrates in soil solutions

In spite of their low concentrations in soil solutions, low–molecular weight organic substances (LMWOS) such as amino acids, sugars, and uronic acids play a major role in the cycles of C and N in soil. With respect to their low concentrations and to possible matrix interferences, their analysis in soil leachates is a challenging task. We established two HPLC (high‐performance liquid chromatography) methods for the parallel determination of amino acids and carbohydrates in soil leachates. The pre‐column derivatization of amino acids with an o‐phthaldialdehyde (OPA) mercaptoethanol solution yields quantitation limits between 0.03 and 0.44 µmol L^–1 and S_D values of <8.3% (n = 9). High‐performance anion‐exchange chromatography (HPAEC) on a Dionex CarboPac PA 20 column with a NaOH acetate gradient combined with pulsed amperometric detection (PAD) was used for the determination of carbohydrates. The calibration curves obtained for 11 carbohydrates showed excellent linearity over the concentration range from 0.02 to 50.0 mg L^–1. Recovery studies revealed good results for all analytes (89%–108%). Interferences from Hg(II) salts and chloroform used for stabilization of the leachates did not occur with both chromatographic methods. The optimized method was successfully used for quantitative determinations of amino acids and carbohydrates in soil leachates.     

3D-X-ray microcomputer tomography and optical coherence tomography as methods for the localization of the blastoderm in the newly laid unincubated chicken egg

The routine culling of the male offspring of hybrid layer type chickens is met with increasing public disapproval for both ethical and legal reasons. Until now practice-oriented methods for reliable sex diagnosis prior to hatch could not be developed. Molecular genetical analysis of blastodermic cells can be used for sex determination in unincubated eggs; however, knowledge of the precise localization of the germinal disc is crucial for the extraction of a carefully directed cell biopsy. In principle, 3D-X-ray micro computed tomography (3D-CT) has been proven a suitable method to localize the germinal disk in the unincubated egg without damaging the egg shell. No negative effects on embryogenesis and hatching rate of irradiated hatching eggs were established. The pictorial representation of the germinal disk using optical coherence tomography (OCT) failed in the unopened egg. The egg shell formed an impenetrable barrier for the currently available measuring method which utilized near infrared (NIR) wavelength regions. After opening the egg shell, the germinal disk could be visualized without any difficulties. In conclusion, technical possibilities for localization of the germinal disk in the unincubated egg already exist, but regarding technical parameters, the procedures have to be adapted to the specific purpose.     

Use of albumin quotient and IgG index to differentiate blood- vs brain-derived proteins in the cerebrospinal fluid of cats with feline infectious peritonitis

BACKGROUND: Inflammation of the central nervous system (CNS) is a frequent condition in cats but etiology often remains unsolved. Routine cerebrospinal fluid (CSF) analysis can be extended through the calculation of the albumin quotient (Q(alb)), a marker of the integrity of the blood-brain barrier (BBB), and IgG index, an estimate of intrathecal IgG synthesis. OBJECTIVES: The purpose of this study was to validate nephelometric methods for CSF protein analysis, and to use the Q(alb) and IgG index to discriminate blood- and brain-derived immunoglobulin fractions in cats with feline infectious peritonitis (FIP). METHODS: Cats presented to our clinic between 2001 and 2005 were included in the study based on clinical and laboratory data and histopathologic findings at necropsy. Cats were grouped as having nonneurologic disease (controls; n=37), brain tumors (n=8), FIP involving the CNS (n=12), and extraneural FIP (n=12). CSF-total protein (TP) was measured and albumin and IgG concentrations were measured in paired CSF/serum samples; Q(alb) and IgG index were calculated. Intraassay and interassay precision of the nephelometric assays were determined using pooled samples. RESULTS: Coefficients of variation for the nephelometric assays ranged from 2.7% to 7.2%. In control cats, CSF-TP concentration ranged from 0.06 to 0.36 g/L, Q(alb) ranged from 0.6 to 5.7 x 10(-3), and IgG index ranged from 0.3 to 0.6. Q(alb) and IgG index were significantly higher in cats with brain tumors and cats with CNS-FIP compared with other groups. Compared with control cats, pleocytosis was evident in 8 of 12 (67%) cats and CSF-TP was increased in 3 of 12 (25%) cats with CNS-FIP. CONCLUSION: Nephelometry is a reliable method for measurement of CSF protein, albumin, and IgG in cats. The Q(alb) and IgG index did not identify a CSF protein pattern specific for BBB dysfunction or intrathecal IgG synthesis in cats with CNS-FIP.     

Effect of Heat Stress on Concentrations of Faecal Cortisol Metabolites in Dairy Cows

The negative impact of heat stress on health and productivity of dairy cows is well known. Heat stress can be quantified with the temperature–humidity index (THI) and is defined as a THI ≥ 72. Additionally, animal welfare is affected in cows living under heat stress conditions. Finding a way to quantify heat stress in dairy cows has been of increasing interest over the past decades. Therefore, the objective of this study was to evaluate concentrations of faecal glucocorticoid metabolites [i.e. 11,17‐dioxoandrostanes (11,17‐DOA)] as an indirect stress parameter in dairy cows without heat stress (DOA 0), with heat stress on a single day (acute heat stress, DOA 1) or with more than a single day of heat stress (chronic heat stress, DOA 2). Cows were housed in five farms under moderate European climates. Two statistical approaches (approach 1 and approach 2) were assessed. Using approach 1, concentrations of faecal 11,17‐DOA were compared among DOA 0, DOA 1 and DOA 2 samples regardless of their origin (i.e. cow, unpaired comparison with a one‐way anova ). Using approach 2, a cow was considered as its own control; that is 11,17‐DOA was treated as a cow‐specific factor and only paired samples were included in the analysis for this approach (paired comparison with t‐tests). In approach 1 (p = 0.006) and approach 2 (p = 0.038), 11,17‐DOA values of cows under acute heat stress were higher compared to those of cows without heat stress. Our results also indicate that acute heat stress has to be considered as a confounder in studies measuring faecal glucocorticoid metabolites in cows to evaluate other stressful situations.     

Intra-tumoral gene delivery of feIL-2, feIFN-gamma and feGM-CSF using magnetofection as a neoadjuvant treatment option for feline fibrosarcomas: a phase-I study

Despite aggressive pre- or postoperative treatment, feline fibrosarcomas have a high relapse rate. In this study, a new treatment option based on immune stimulation by intra-tumoral delivery of three feline cytokine genes was performed. The objective of this phase-I dose-escalation study was to determine a safe dose for further evaluation in a subsequent phase-II trial. Twenty-five client-owned cats with clinical diagnosis of fibrosarcoma - primary tumours as well as recurrences - entered the study. Four increasing doses of plasmids coding for feIL-2, feIFN-gamma or feGM-CSF, respectively, were previously defined. In groups I, II, III and IV these doses were 15, 50, 150 and 450 microg per plasmid and a corresponding amount of magnetic nanoparticles. Two preoperative intra-tumoral injections of the magnetic DNA solution were followed by magnetofection. A group of four control cats received only surgical treatment. Side effects were registered and graded according to the VCOG-CTCAE scale and correlated to treatment. Statistical analyses included one-way anova, post hoc and Kruskal-Wallis tests. ELISA tests detecting plasma feIFN-gamma and plasma feGM-CSF were performed. One cat out of group IV (450 microg per plasmid) showed adverse events probably related to gene delivery. As these side effects were self-limiting and occurred only in one of eight cats in group IV, this dose was determined to be well tolerable. Altogether six cats developed local recurrences during a 1-year observation period. Four of these cats had been treated with dose IV. Regarding these observations, a subsequent phase-II trial including a representative amount of cats should be tested for the efficacy of dose IV as well as dose III.