New host record of five Flavobacterium species associated with tropical fresh water farmed fishes from North India

Abstract Yellow pigmented, filamentous, Gram-negative bacteria belonging to genus Flavobacterium are commonly associated with infections in stressed fish. In this study, inter-species diversity of Flavobacterium was studied in apparently healthy freshwater farmed fishes. For this, ninety one yellow pigmented bacteria were isolated from skin and gill samples (n = 38) of three farmed fish species i.e. Labeo rohita, Catla catla and Cyprinus carpio. Among them, only twelve bacterial isolates (13.18%) were identified as Flavobacterium spp. on the basis of morphological, biochemical tests, partial 16S rDNA gene sequencing and phylogenetic analysis. On the basis of 16S rDNA gene sequencing, all the 12 isolates were 97.6-100% similar to six different formally described species of genus Flavobacterium. The 16S rDNA based phylogenetic analysis grouped these strains into six different clades. Of the 12 isolates, six strains (Fl9S1-6) grouped with F. suncheonense, two strains (Fl6I2, Fl6I3) with F. indicum and the rest four strains (Fl1A1, Fl2G1, Fl3H1 and Fl10T1) clustered with F. aquaticum, F. granuli, F. hercynium and F. terrae, respectively. None of these species except, F. hercynium were previously reported from fish. All the isolated Flavobacterium species possessed the ability of adhesion and biofilm formation to colonize the external surface of healthy fish. The present study is the first record of tropical freshwater farmed fishes as hosts to five environmentally associated species of the Flavobacterium.

Composition of Extracellular Polymeric Substances (EPS) produced by Flavobacterium columnare isolated from tropical fish in Brazil

Thirty nine isolates of Flavobacterium columnare from Brazilian fish farms had their carbohydrate composition of EPS evaluated by high efficiency liquid chromatography, using the phenol-sulfuric acid method of EPS. The occurrence of capsules on F. columnare cells was not directly related to biofilm formation, and the predominant monosaccharide is glucose.

Studies on flavobacterium columnare isolated from fresh water fish

Flavobacterium columnre is one of the most important bacterial pathogens of wild, farmed and ornamental fish in warm, and also in fresh and sea water causing columnaris disease. In the present study one hundred and forty Nile tilapia were collected from different localities of farms from El-Behera governorate and Kafr El-sheikh provinces. The clinical signs of the disease were detected and the pathological lesions were recorded. Bacteriological examination of samples of naturally infected fish with Flavobacterium columnare revealed that the characteristics colonies on cytophaga agar media. Sixty seven suspected isolates were isolated and biochemically identified by API-20 E system. The distribution of the gained isolates among organs denoted 33 isolates from gills [49.25%], 20 isolates from kidneys [29-85%], 12 isolates from deep layer of dorsal musculature [17.91%], one isolate from both spleen and intestine [1.49%], but it was not isolated from the liver. The pathogencity experiment indicated wide variation among Flavobacterium columnare isolates. Isolate number 10 was the most virulent isolate as it caused 100% mortality two weeks post injection followed by isolate number 7, 3 caused 45, 25% mortality. The clinical signs and the post mortem examination revealed the same signs and lesions of natural infection with Flavobacterium columnare beside severe oedematous swelling at site of injection. The median lethal dose [LD50] of the highly virulent isolate was 10[5] viable bacterial cells /ml. Stress factors, temperature, PH, unionized ammonia and dissolved oxygen affects on the pathogencity of the organism. In vitro antibiotic sensitivity test of 5 isolates revealed that the most effective drug Cefotaxime [60%]. While the less effective one, were Neomycin, Norfloxacin and Rifampcin [100%] then Doxycyclin, Cephalexin, Florfenicel, Gentamycin and Marbocyl [80%] then Cefalor, Tetracycline and Ciprofloxacin [60%]. On the other hand, Erthromycin, Ampicillin [80%] had no effect

Isolation and characterization of Flavobacterium columnare (Bernardet et al. 2002) from four tropical fish species in Brazil / Isolamento e caracterização de Flavobacterium columnare de quatro espécies de peixes tropicais no Brasil

Flavobacterium columnare is the causative agent of columnaris disease in freshwater fish, implicated in skin and gill disease, often causing high mortality. The aim of this study was the isolation and characterization of Flavobacterium columnare in tropical fish in Brazil. Piracanjuba (Brycon orbignyanus), pacu (Piaractus mesopotamicus), tambaqui (Colossoma macropomum) and cascudo (Hypostomus plecostomus) were examined for external lesions showing signs of colunmaris disease such as greyish white spots, especially on the head, dorsal part and caudal fin of the fish. The sampling comprised 50 samples representing four different fish species selected for study. Samples for culture were obtained by skin and kidney scrapes with a sterile cotton swabs of columnaris disease fish and streaked onto Carlson and Pacha (1968) artificial culture medium (broth and solid) which were used for isolation. The strains in the liquid medium were Gram negative, long, filamentous, exhibited flexing movements (gliding motility), contained a large number of long slender bacteria and gathered into ‘columns'. Strains on the agar produced yellow-pale colonies, rather small, flat that had rhizoid edges. A total of four Flavobacterium columnare were isolated: 01 Brycon orbignyanus strain, 01 Piaractus mesopotamicus strain, 01 Colossoma macropomum strain, and 01 Hypostomus plecostomus strain. Biochemical characterization, with its absorption of Congo red dye, production of flexirubin-type pigments, H2S production and reduction of nitrates proved that the isolate could be classified as Flavobacterium columnare.

Flavobacterium columnare é o agente etiológico da columnariose em peixes de água doce, ocasionando enfermidade na pele e nas brânquias, provocando freqüentemente um grande número de mortalidade. O objetivo deste estudo foi o isolamento e a caracterização de Flavobacterium columnare em peixes tropicais no Brasil. Piracanjuba (Brycon orbignyanus), pacu (Piaractus mesopotamicus), tambaqui (Colossoma macropomum) e cascudo (Hypostomus plecostomus) foram examinados externamente com relação a sinais característicos de columnariose, como manchas acinzentadas na cabeça, região dorsal e pedúnculo caudal dos peixes. A amostragem compreendeu a coleta de 50 exemplares de peixes, representando as quatro diferentes espécies escolhidas para este estudo. Amostras para o isolamento foram obtidas através de raspado com swab estéril das lesões e do rim dos peixes clinicamente diagnosticados como acometidos por columnarios e imediatamente semeados em meios de culturas artificiais (líquido e sólido) próprios para o estudo de Flavobacterium segundo Carlson e Pacha (1968). No meio líquido, houve o desenvolvimento de microrganismos que observados em gota pendente apresentaram a forma de bacilos finos, longos, móveis por deslizamento. Através da coloração de Gram, apresentaram morfologia de bacilos finos, Gram negativos, agrupados em colunas. Em meio sólido, as colônias eram pequenas, cinza-amareladas, com borda em forma de raiz. No total, foram obtidos quatro isolamentos: 01 cepa de Brycon orbignyanus; 01 cepa de Piaractus mesopotamicus; 01 cepa de Colossoma macropomum; e 01 cepa de Hypostomus plecostomus. A caracterização bioquímica das amostras, como absorção do vermelho Congo, produção de flexirrubina, produção de H2S e redução do nitrato, sugere que os isolamentos poderiam ser classificados como Flavobacterium columnare.

Prevalence of nonfermenters in clinical specimens.

One hundred and thirty three non fermenting gram negative bacilli isolated out of 625 different clinical specimens were identified and characterised. Samples were exudate from chronic suppractive otits media (341), diabetic foot (117) wound (116) and blood (51). Of these isolates Pseudomonas aeruginosa 105(78.94%) predominated followed by Acinetobacter sp 8 [6.1%], Pseudomonas putrifaciens 6(4.5%), Flavobacterium sp 6(4.5%), Xanthomonas maltophilia 5(3.75%), Alkaligenes sp 3 (2.25). 31 (23.30%) were resistant to commonly used antibiotics. Amikacin 85 (63.90%) was found to be more effective than fluoroquinolones (27.8-48.12%).

An unusual case of Flavobacterium meningosepticum pneumonia in an immunocompromised patient.

An unusual case of Flavobacterium meningosepticum pneumonia in an immunocompromised patient is reported. The necessity to recognize this opportunistic pathogen and its implication is discussed.

Preparation and purification of Flavobacterium heparinum chondroitinases AC and B by hydrophobic interaction chromatography

Flavobacterium heparinum is a soil bacterium that produces several mucopolysaccharidases such as heparinase, heparitinases I and II, and chondroitinases AC, B, C and ABC. The purpose of the present study was to optimize the preparation of F. heparinum chondroitinases, which are very useful tools for the identification and structural characterization of chondroitin and dermatan sulfates. We observed that during the routine procedure for cell disruption (ultrasound, 100 kHz, 5 min) some of the chondroitinase B activity was lost. Using milder conditions (2 min), most of the chondroitinase B and AC protein was solubilized and the enzyme activities were preserved. Tryptic soy broth without glucose was the best culture medium both for bacterial growth and enzyme induction. Chondroitinases AC and B were separated from each other and also from glucuronidases and sulfatases by hydrophobic interaction chromatography on HP Phenyl-Sepharose. A rapid method for screening of the column fractions was also developed based on the metachromatic shift of the color of dimethylmethylene blue

Frequency of nonfermentative gram-negative Bacilli isolated from clinical materials of pacients at Universidade Federal do Ceará Hospital Complex - Brazil

Among one thousand eight hundred and thirty-four Gram-negative bacilli, isolated at UniversidadeFederal do Ceará hospital complex-Brazil, from January 1995 to February 1996, 456 (24,8 per cent) were Nonfermentative Gram-Negative Bacilli (NFGNB). This study reports their identification to the species level and their frequency as well. Thirteen genera and thirty species were identified and Pseudomonas aeruginosa was the most frequent species (69,95 per cent), followed by Acinetobacter baumannii (5,48 per cent) and by Acinetobacter lwoffii (3,95 per cent). Among the identified P.aeruginosa strains, (94,1 per cent) produced pigment but (7,9 per cent) of them produced pigment only after being cultivated several times. The frequency of the most species was similar to that reported in the literature

Sphingobacterium multivorum septicemia: a case report.

A 47-year-old man presented with a history of fever, chills and weight loss for 3 months. He had been treated for diabetes mellitus during the past 3 years. He developed high fever with abnormal liver function tests. Both Widal and Weil-Felix reactions were negative with normal roentgenogram of the chest. His anti-HIV tests were positive. The cultures from the blood and sputum yielded pure Sphingobacterium multivorum sensitive to sulfamethoxazole-trimethoprim, chloramphenicol, tetracycline, cefotaxime, ceftazidine and ceftriaxone. On the next day, the patient developed signs and symptoms of meningitis with the CSF containing chronic and acute inflammatory cells but revealed no growth on culture. The patient was treated with a combination of ceftriazone and trimethoprim-sulfamethoxazole but he died on the 6th day after admission. This patient was the fifth reported case infected with S.multivorum. It illustrates that this potentially pathogenic organism can cause septicemia in an immunodeficient patient.