The Role of Host Species in Experimental Ferlavirus Infection: Comparison of a Single Strain in Ball Pythons (Python regius) and Corn Snakes (Pantherophis guttatus).

Ferlaviruses are a cause of respiratory disease in snakes. Four genogroups (A, B, C, and tortoise) have been described. Disease development is believed to depend on virus, host, and environment-specific factors. There is evidence of transmission of individual strains between genera and families of reptiles. A genogroup B virus previously used in a transmission study with corn snakes (Pantherophis guttatus) was applied intratracheally in ball pythons (Python regius) using the same protocol as for the corn snakes. Ball pythons became infected, with initial mild clinical signs noted four days post infection (p.i.), and the virus was detected first in the lungs on day 4 and spread to the intestine, pancreas, kidney and brain. Hematology showed an increase in circulating lymphocytes which peaked on day 28 p.i. Antibodies were detected beginning on day 16 and increased steadily to the end of the study. In comparison to corn snakes, ball pythons exhibited milder clinical signs and pathological changes, faster development of and higher antibody titers, and a hematological reaction dominated by lymphocytosis in contrast to heterophilia in corn snakes. These differences in host reaction to infection are important to understand ferlavirus epidemiology as well as for clinical medicine and diagnostic testing.

A distinct variant of the SzM protein of Streptococcus equi subsp. zooepidemicus recruits C1q independent of IgG binding and inhibits activation of the classical complement pathway.

Streptococcus equi subsp. zooepidemicus (SEZ) is a major equine pathogen that causes pneumonia, abortion, and polyarthritis. It can also cause invasive infections in humans. SEZ expresses the M-like protein SzM, which recruits host proteins such as fibrinogen to the bacterial surface. Equine SEZ strain C2, which binds only comparably low amounts of human fibrinogen in comparison to human SEZ strain C33, was previously shown to proliferate in equine and human blood. As the expression of SzM_C2 was necessary for survival in blood, this study investigated the working hypothesis that SzM_C2 inhibits complement activation through a mechanism other than fibrinogen and non-immune immunoglobulin binding. Loss-of-function experiments showed that SEZ C2, but not C33, binds C1q via SzM in IgG-free human plasma. Furthermore, SzM C2 expression is necessary for recruiting purified human or equine C1q to the bacterial surface. Flow cytometry analysis demonstrated that SzM expression in SEZ C2 is crucial for the significant reduction of C3b labelling in human plasma. Addition of human plasma to immobilized rSzM_C2 and immobilized aggregated IgG led to binding of C1q, but only the latter activated the complement system, as shown by the detection of C4 deposition. Complement activation induced by aggregated IgG was significantly reduced if human plasma was pre-incubated with rSzM_C2. Furthermore, rSzM_C2, but not rSzM_C33, inhibited the activation of the classical complement pathway in human plasma, as determined in an erythrocyte lysis experiment. In conclusion, the immunoglobulin-independent binding of C1q to SzM_C2 is associated with complement inhibition.

Molecular epidemiology of Trichophyton quinckeanum - a zoophilic dermatophyte on the rise.

BACKGROUND: Formerly only referred to as a subspecies (T. mentagrophytes var. quinckeanum), T. quinckeanum once again constitutes a distinct species according to the updated taxonomy of dermatophytes. PATIENTS AND METHODS: During routine diagnostic tests conducted at the Mycology Laboratory, Mölbis, Germany, between 11/2013 to 1/2017 (three years and three months), all specimens sent in were examined for T. quinckeanum. Molecular biology methods employed included: 1) DNA hybridization (PCR ELISA), 2) gene sequencing of the ITS region and TEF-1α, and 3) in some cases, MALDI-TOF mass spectrometry. RESULTS: Overall, 62 strains of T. quinckeanum were found. Sixty-eight percent of patients were female; 43 % were children and adolescents (≤ 19 years of age). Cats were a frequent source of infection. Sequencing of all 62 strains revealed a concordance of 100 % with T. quinckeanum sequences contained in the NCBI database. Isolates analyzed by MALDI-TOF mass spectrometry showed specific spectra. CONCLUSIONS: In Germany, the zoophilic dermatophyte T. quinckeanum currently appears to be more prevalent than expected. T. quinckeanum strains were isolated from children and adults with dermatomycosis and tinea capitis. Sources of infection with T. quinckeanum include small rodents (mice), horses, and - remarkably commonly -  cats. Given that unequivocal morphological identification of this dermatophyte is not always possible, molecular methods have to be employed in the diagnosis.

Acute phase proteins as local biomarkers of respiratory infection in calves.

BACKGROUND: Cumulating reports suggest that acute phase proteins (APPs) do not only play a role as systemic inflammatory mediators, but are also expressed in different tissues as local reaction to inflammatory stimuli. The present study aimed to evaluate presence and changes in luminal lung concentrations of the APPs haptoglobin (Hp), lipopolysaccharide binding protein (LBP), C-reactive protein (CRP), and lactoferrin (Lf) in calves with an acute respiratory disease experimentally induced by Chlamydia (C.) psittaci. RESULTS: Intra-bronchial inoculation of the pathogen resulted in a consistent respiratory illness. In venous blood of the infected calves (n = 13), concentrations of plasma proteins and serum LBP were assessed (i) before exposure and (ii) 8 times within 14 days after inoculation (dpi). Increasing clinical illness correlated significantly with increasing LBP-and decreasing albumin concentrations in blood, both verifying a systemic acute phase response. Broncho-alveolar lavage fluid (BALF) was obtained from all 13 calves experimentally infected with C. psittaci at 4, 9 and 14 dpi, and from 6 uninfected healthy calves. Concentrations of bovine serum albumin (BSA), Hp, LBP, CRP and Lf in BALF were determined by ELISA. In infected animals, absolute concentrations of LBP and Hp in BALF correlated significantly with the respiratory score. The quotient [LBP]/[BSA] in BALF peaked significantly in acutely infected animals (4 dpi), showed a time-dependent decrease during the recovery phase (9-14 dpi), and was significantly higher compared to healthy controls. Concentrations of Hp and Lf in BALF as well as [Hp]/[BSA]--and [Lf]/[BSA]-quotients decreased during the study in infected animals, but were never higher than in healthy controls. CRP concentrations and [CRP]/[BSA]-quotient did not express significant differences between infected and healthy animals or during the course of infection. CONCLUSION: In conclusion, absolute concentrations of LBP in blood and BALF as well as the quotient [LBP]/[BSA] in BALF perfectly paralleled the clinical course of respiratory illness after infection. Beside LBP, the suitability of Hp and Lf as local biomarkers of respiratory infections in cattle and their role in the local response to pathogens is worth further investigation, while CRP does not seem to play a role in local defense mechanisms of the bovine lung.

Development of intestinal microflora and occurrence of diarrhoea in sucking foals: effects of Bacillus cereus var. toyoi supplementation.

BACKGROUND: Almost all foals develop transient diarrhoea within the first weeks of life. Studies indicated different viral, bacterial, and parasitic causes, such as rotavirus, Clostridium perfringens, Escherichia coli, and Cryptosporidium are discussed. But little is known about the development of intestinal microflora in foals. The present study investigated whether the supplementation with Bacillus cereus var. toyoi would modify the developing intestinal microflora and consequently reduce diarrhoea in foals. From birth, the foals were randomly assigned to three treatment groups: placebo (10 mL isotonic NaCl, n = 8), low dosage (LD; 5 × 10(8) cfu B. cereus var. toyoi, n = 7) and high dosage (HD; 2 × 10(9) cfu B. cereus var. toyoi, n = 10). Treatment groups were supplemented orally once a day for 58 days. Faeces scoring and sampling were performed within the first 24 h after birth and on day 9, 16, 23, 30, 44, 58 of the foal's life and also on the first day of diarrhoea. Culture-plate methods were used to analyse the bacterial microflora. RESULTS: Eighty-eight per cent of the foals developed diarrhoea (placebo 7/8, LD 5/7, HD 10/10) during the first 58 days of life. Bacillus cereus var. toyoi supplementation had no effect on bacterial microflora. Clostridium perfringens and enterobacteria were equally prevalent in foals with diarrhoea and those who were not afflicted. CONCLUSIONS: We conclude that the supplementation of B. cereus var. toyoi had no effect on the occurrence of diarrhoea and health status in the foals.

Enrofloxacin and macrolides alone or in combination with rifampicin as antimicrobial treatment in a bovine model of acute Chlamydia psittaci infection.

Chlamydia psittaci is a zoonotic bacterium with a wide host range that can cause respiratory disease in humans and cattle. In the present study, effects of treatment with macrolides and quinolones applied alone or in combination with rifampicin were tested in a previously established bovine model of respiratory C. psittaci infection. Fifty animals were inoculated intrabronchially at the age of 6-8 weeks. Seven served as untreated controls, the others were assigned to seven treatment groups: (i) rifampicin, (ii) enrofloxacin, (iii) enrofloxacin + rifampicin, (iv) azithromycin, (v) azithromycin + rifampicin, (vi) erythromycin, and (vii) erythromycin + rifampicin. Treatment started 30 hours after inoculation and continued until 14 days after inoculation (dpi), when all animals were necropsied. The infection was successful in all animals and sufficient antibiotic levels were detected in blood plasma and tissue of the treated animals. Reisolation of the pathogen was achieved more often from untreated animals than from other groups. Nevertheless, pathogen detection by PCR was possible to the same extent in all animals and there were no significant differences between treated and untreated animals in terms of local (i.e., cell count and differentiation of BALF-cells) and systemic inflammation (i.e. white blood cells and concentration of acute phase protein LBP), clinical signs, and pathological findings at necropsy. Regardless of the reduced reisolation rate in treated animals, the treatment of experimentally induced respiratory C. psittaci infection with enrofloxacin, azithromycin or erythromycin alone or in combination with rifampicin was without obvious benefit for the host, since no significant differences in clinical and pathological findings or inflammatory parameters were detected and all animals recovered clinically within two weeks.

A bovine model of a respiratory Parachlamydia acanthamoebae infection.

The aim of this study was to evaluate the pathogenicity of Parachlamydia (P.) acanthamoebae as a potential agent of lower respiratory tract disease in a bovine model of induced lung infection. Intrabronchial inoculation with P. acanthamoebae was performed in healthy calves aged 2-3 months using two challenge doses: 10(8) and 10(10) bacteria per animal. Controls received 10(8) heat-inactivated bacteria. Challenge with 10(8) viable Parachlamydia resulted in a mild degree of general indisposition, whereas 10(10) bacteria induced a more severe respiratory illness becoming apparent 1-2 days post inoculation (dpi), affecting 9/9 (100%) animals and lasting for 6 days. The extent of macroscopic pulmonary lesions was as high as 6.6 (6.0)% [median (range)] of lung tissue at 2-4 dpi and correlated with parachlamydial genomic copy numbers detected by PCR, and with bacterial load estimated by immunohistochemistry in lung tissue. Clinical outcome, acute phase reactants, pathological findings and bacterial load exhibited an initial dose-dependent effect on severity. Animals fully recovered from clinical signs of respiratory disease within 5 days. The bovine lung was shown to be moderately susceptible to P. acanthamoebae, exhibiting a transient pneumonic inflammation after intrabronchial challenge. Further studies are warranted to determine the precise pathophysiologic pathways of host-pathogen interaction.

Evaluation of antimicrobial treatment in a bovine model of acute Chlamydia psittaci infection: tetracycline versus tetracycline plus rifampicin.

Antimicrobial treatment of chlamydial infections is known to be of limited efficacy. In this study, effects of doxycycline (D), usually the drug of choice, were compared with the combined therapy of doxycycline and rifampicin (R) in a bovine model of respiratory Chlamydia psittaci infection. After intrabronchial inoculation of the pathogen, 30 animals were assigned to five groups (n = 6 per group): untreated controls, monotherapy with D (5 mg kg(-1)day(-1) or 10 mg kg(-1)day(-1)), and combination therapy of D and R (600 mg day(-1)). Treatment continued until day 14 post inoculation (d.p.i.). Clinical signs, inflammatory markers, and pathological findings confirmed successful infection in all animals. Reisolation of the pathogen was possible in 4/6 untreated animals and in 4/12 animals treated with D alone until 4 d.p.i., but in none of the calves of the two D + R groups. Pathogen detection was possible in all animals without significant differences among groups. Severity of disease and time course of its resolution, assessed by clinical and pathological findings as well as inflammatory parameters, were not significantly different between untreated controls and calves receiving D alone or in combination with R. Regardless of the treatment regimen, all groups recovered clinically and cleared the infection within 2 weeks.

Intestinal blood flow in patients with chronic heart failure: a link with bacterial growth, gastrointestinal symptoms, and cachexia.

BACKGROUND: Blood flow in the intestinal arteries is reduced in patients with stable heart failure (HF) and relates to gastrointestinal (GI) symptoms and cardiac cachexia. OBJECTIVES: The aims of this study were to measure arterial intestinal blood flow and assess its role in juxtamucosal bacterial growth, GI symptoms, and cachexia in patients with HF. METHODS: A total of 65 patients and 25 controls were investigated. Twelve patients were cachectic. Intestinal blood flow and bowel wall thickness were measured using ultrasound. GI symptoms were documented. Bacteria in stool and juxtamucosal bacteria on biopsies taken during sigmoidoscopy were studied in a subgroup by fluorescence in situ hybridization. Serum lipopolysaccharide antibodies were measured. RESULTS: Patients showed 30% to 43% reduced mean systolic blood flow in the superior and inferior mesenteric arteries and celiac trunk (CT) compared with controls (p < 0.007 for all). Cachectic patients had the lowest blood flow (p < 0.002). Lower blood flow in the superior mesenteric artery and CT was correlated with HF severity (p < 0.04 for all). Patients had more feelings of repletion, flatulence, intestinal murmurs, and burping (p < 0.04). Burping and nausea or vomiting were most severe in patients with cachexia (p < 0.05). Patients with lower CT blood flow had more abdominal discomfort and immunoglobulin A-antilipopolysaccharide (r = 0.76, p < 0.02). Antilipopolysaccharide response was correlated with increased growth of juxtamucosal but not stool bacteria. Patients with intestinal murmurs had greater bowel wall thickness of the sigmoid and descending colon, suggestive of edema contributing to GI symptoms (p < 0.05). In multivariate regression analysis, lower blood flow in the superior mesenteric artery, CT (p < 0.04), and inferior mesenteric artery (p = 0.056) was correlated with the presence of cardiac cachexia. CONCLUSIONS: Intestinal blood flow is reduced in patients with HF. This may contribute to juxtamucosal bacterial growth and GI symptoms in patients with advanced HF complicated by cachexia.

Trichophyton species of Arthroderma benhamiae - a new infectious agent in dermatology.

In Germany, infections due to the zoophilic dermatophyte Trichophyton (T.) species of Arthroderma benhamiae are being more frequently diagnosed. The source of infection of this emerging pathogen overlaps with that of the zoophilic species T. interdigitale. The most common source are guinea pigs. T. species of Arthroderma benhamiae causes inflammatory dermatophytosis in children and adolescents. In addition to tinea capitis, it may cause both tinea corporis, tinea manus and frequently tinea faciei. In Germany, T. species of Arthroderma benhamiae is a frequent zoophilic dermatophyte, which in regions is probably more frequent than Microsporum canis. The mycological identification of the isolates with their yellow stained colonies is based on their macroscopic and microscopic features. However, some exhibit colony features consistent with those of T. interdigitale. These strains only can be identified unambiguously by means of molecular techniques. Using detection methods such as PCR-ELISA or real-time PCR, the dermatophyte can be identified directly from clinical material. Sequencing of the internal transcribed spacer region (ITS) of the ribosomal DNA has been approved as culture confirmation test for T. species of Arthroderma benhamiae. In addition, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS) is useful. Widespread dermatophytosis due to T. species of Arthroderma benhamiae, in particular of tinea capitis, requires oral antifungal agents. Terbinafine is most effective, alternatives are fluconazole and itraconazole.

Dose-dependent effects of Chlamydia psittaci infection on pulmonary gas exchange, innate immunity and acute-phase reaction in a bovine respiratory model.

The respiratory pathogen Chlamydia psittaci naturally occurs in bovine herds and was recently shown to impair calf health in a dose-dependent manner. The aim of this study was to determine whether the functional consequences and immunological reactions of infection were dose related by quantifying the consequences of acute respiratory chlamydial infection on respiratory signs, disturbances of pulmonary gas exchange, response of the innate immune system, and acute-phase reaction. Fourteen calves were challenged intrabronchially with different C. psittaci doses (from 10(6) to 10(9)inclusion-forming units (ifu) per animal). Ten controls received either UV-inactivated chlamydiae or cell culture medium. Compared to the controls, all animals challenged with live C. psittaci developed hypoxaemia linked to reduced haemoglobin oxygen saturation, increased alveolar-arterial oxygen partial pressure difference (A-aO2) and pulmonary shunt, with symptoms following a dose-dependent pattern. Increases in lipopolysaccharide-binding protein (LBP) and leukocytes were also dose-dependent and accompanied by a regenerative left shift in neutrophil granulocytes. With the exception of LBP, which reflected the load of chlamydial cell components in the host, pathophysiological reactions were only detected in calves challenged with viable chlamydiae. These results indicate that the pathophysiological consequences of respiratory C. psittaci infections are strongly dependent on the challenge dose of chlamydiae. For further studies, challenge doses between 10(6) and 10(8)ifu/calf are recommended.

Altered intestinal function in patients with chronic heart failure.

OBJECTIVES: We evaluated morphology and function of the gut in patients with chronic heart failure (CHF). BACKGROUND: Intestinal translocation of bacterial endotoxin may contribute to the inflammatory state observed in patients with CHF. The morphology and function of the gut may be abnormal. METHODS: We studied 22 patients with CHF (age 67 +/- 2 years, left ventricular ejection fraction [LVEF] 31 +/- 1%, New York Heart Association functional class 2.3 +/- 0.1, peak VO2 15.0 +/- 1.0 ml/kg/min) and 22 control subjects (62 +/- 1 years, LVEF 68 +/- 2%, peak VO2 24.7 +/- 1.3 ml/kg/min). Bowel wall thickness was assessed by transcutaneous sonography, small intestinal permeability by the lactulose-mannitol test, passive carrier-mediated transport by D-xylose test, large intestinal permeability by sucralose test (5- and 26-h urine collection, high-performance liquid chromatography), and mucosal bacterial biofilm by fluorescence in situ hybridization in biopsies taken during sigmoidoscopy. RESULTS: Chronic heart failure patients, compared with control patients, showed increased bowel wall thickness in the terminal ileum (1.48 +/- 0.16 mm vs. 1.04 +/- 0.08 mm), ascending colon (2.32 +/- 0.18 mm vs. 1.31 +/- 0.14 mm), transverse colon (2.19 +/- 0.20 vs. 1.27 +/- 0.08 mm), descending colon (2.59 +/- 0.18 mm vs. 1.43 +/- 0.13 mm), and sigmoid (2.97 +/- 0.27 mm vs. 1.64 +/- 0.14 mm) (all p < 0.01). Chronic heart failure patients had a 35% increase of small intestinal permeability (lactulose/mannitol ratio: 0.023 +/- 0.001 vs. 0.017 +/- 0.001, p = 0.006), a 210% increase of large intestinal permeability (sucralose excretion: 0.62 +/- 0.17% vs. 0.20 +/- 0.06%, p = 0.03), and a 29% decrease of D-xylose absorption, indicating bowel ischemia (26.7 +/- 3.0% vs. 37.4 +/- 1.4%, p = 0.003). Higher concentrations of adherent bacteria were found within mucus of CHF patients compared with control subjects (p = 0.007). CONCLUSIONS: Chronic heart failure is a multisystem disorder in which intestinal morphology, permeability, and absorption are modified. Increased intestinal permeability and an augmented bacterial biofilm may contribute to the origin of both chronic inflammation and malnutrition.

Microbial and immunological responses relative to high-altitude exposure in mountaineers.

PURPOSE: High-altitude exposure is often associated with gastrointestinal disorders, inflammation, and an increased risk of infection. We suspected microbial and immunological responses to high-altitude exposure in mountaineers resulting from changes in the balance of the intestinal microflora. METHODS: We investigated fecal samples and serum of seven mountaineers who took part in a 47-d German expedition to the Nepalese Himalayas in 2002, for microbial response by changes in different fecal bacterial population groups (fluorescent in situ hybridization), immune response by serum levels of IgG-, IgM-, and IgA anti-LPS (E. coli J5), and inflammatory response by serum levels of C-reactive protein (CRP) (ELISA). In addition, measurements of body temperature, pulse rate, arterial oxygen saturation, and questionnaire (Lake Louise score for altitude illness) were performed. RESULTS: The data indicate a distinct alteration in the composition of the fecal microflora relative to high-altitude exposure above 5000 m. Bifidobacteria and species belonging to the Atopobium, Coriobacterium, and Eggerthella lenta group decreased, whereas potential pathogenic bacteria of the gamma subdivision of Proteobacteria and specific Enterobacteriaceae such as Escherichia coli increased. Possible endotoxemia resulting from the increase of the latter Gram-negative bacteria was indirectly indicated by the reduction in serum levels of IgM- and/or IgA anti-LPS. CRP was elevated relative to high-altitude exposure. The Lake Louise score correlated with the changes in CRP, IgA-, and IgM anti-LPS but did not correlate with the bacterial alterations. CONCLUSION: Changes in the composition of intestinal microbiota may be associated with indicators of an immunological challenge and may result in an increased health risk of mountaineers during exposure to very high altitude.