A new selective medium for the isolation of Listeria monocytogenes.

A new selective medium for the isolation of Listeria monocytogenes (Lm), is described. The medium contained propolis, nalidixic acid, polymyxin B and rivanol as selective substances. The new medium (propolis-agar) was compared with two other selective media and one nonselective medium. No inhibitory effect was found on the 6 strains of Lm tested, and Lm was easily isolated from a mixture of Lm and contaminating bacteria. The selective effect was better than for the two other selective media tested.     

The marked increase of Listeria monocytogenes isolation from contents of swine cecum

The actual prevalence of Listeria monocytogenes from contents of swine cecum was investigated. The efficiency of Listeria enrichment broth (LEB) for isolation was examined by the recovery of artificially inoculated L. monocytogenes in contents of swine cecum. The numbers of organisms did not increase after 48 h incubation, but increased when the rapid decrease in pH of the LEB was adjusted. Between 1991 and 1993, 250 contents of swine cecum were examined for the prevalence of L. monocytogenes using LEB enrichment, either with or without pH adjustment. L. monocytogenes was isolated from 74 samples in 1993 with pH adjustment, however, no organisms were isolated in 1991 and 1992. It was suggested that the marked rise of the L. monocytogenes isolation was due to the spread of the organism among swine. Furthermore, 67 out of the 74 isolates were identified as 1/2c by serotyping. The serovar 1/2c strains showed genetic diversity by random amplified polymorphic DNA.     

Selective effect of propolis in the isolation of Listeria monocytogenes (author's transl)]

Propolis is a substance produced by honeybees. It is inhibitory to some bacteria species, mainly Gram-positive bacteria, but less inhibitory to Listeria monocytogenes (L.m) than to the other Gram-positive bacteria tested. In order to obtain selective growth of L.m. from contaminated samples, the effect of propolis in plating media and broths on various strains of bacteria was examined. Table I shows the effect of increasing concentrations of propolis in tryptose-agar (TA). L.m. tolerated higher concentrations of propolis than Streptococcus viridans and Staphylococcus aureus. L.m. grew well in tryptosebroth (TB) that contained 0.15 mg propolis pr. ml medium, while Streptococcus viridans and Streptococcus agalactiae were completely inhibited as seen in Table II. Table III shows that when serum was added to the agar, the inhibitory effect was reduced. It can also be seen that Gram-negative bacteria grew quite well on media that contained 0.19 mg propolis pr. ml. To reduce the growth of Gram-negative bacteria, nalidixic acid was added to the medium. Table IV illustrates growth of various species of bacteria in tryptosephosphatebroth (TFB) with or without propolis and nalidixic acid. Most of the strains tested were inhibited, but Pseudomonas aeruginosa and to some extent faecal streptococci were able to grow in the medium that contained the selective substances. As a conclusion it seems that propolis may be a valuable additive to a medium for the selective isolation of L.m.     

Influence of bacteriocin-like substance, generation times, and genetic profiles of Listeria innocua on the isolation of Listeria monocytogenes

Inhibition of isolation of Listeria monocytogenes by bacteriocin-like substance (BLS)-producing Listeria innocua after enrichment culture was investigated. When 26 L. monocytogenes strains were examined in combination with eight L. innocua strains using the spot on lawn method, 52/208 (25.0%) combinations showed the growth inhibition of L. monocytogenes. When two Listeria species were cultured simultaneously in selective enrichment broth, inhibition of isolation of L. monocytogenes was observed in 12/52 of the combinations at 24h (23.1%), in 24/52 at 48h (46.2%) and in 30/52 (57.7%) after 7 days of incubation. The randomly amplified polymorphic DNA profiles showed no interstrain similarities between either strains of the BLS-producing L. innocua or the BLS-sensitive L. monocytogenes strains. Therefore inhibition by BLS-producing L. innocua of isolation of L. monocytogenes after enrichment culture is unlikely to be dependent upon a particular genetic profile.     

原代牛肝细胞分离和培养方法的建立

采用胶原酶灌注消化法,对来自新生小牛血清制备厂的牛肝脏尾状突材料进行肝细胞分离,用台盼蓝拒染法测定总细胞数和肝细胞即时存活率,以每孔1×106个肝细胞的量接种于牛尾胶原包被的6孔细胞培养板,进行单层培养细胞形态学观察和培养基上清液LDH活性测定.结果显示,每头小牛肝脏尾状突分离获得的细胞产量为(3.558±0.096)×108个,肝细胞即时存活率为(84.46±3.56)%,培养24～72 h的肝细胞生长状态最好,适合用于有关肝细胞代谢、中毒、基因表达的研究.     

Comparison of five culture methods for Salmonella isolation from swine fecal samples of known infection status

The current study was conducted to evaluate 5 bacteriologic culture methods (methods 1, 2, 3, 4, and 5) for recovery of Salmonella enterica from swine feces, both for sensitivity of detection (ability to recover Salmonella from a positive sample) and for specificity (not to inadvertently identify an organism as a Salmonella species in a negative sample). Fifty-six negative samples and 46 positive samples were processed using each of the 5 methods, which differed primarily in the combinations of enrichment media used. All negative samples were negative for Salmonella when cultured by all 5 methods (100% specificity). Two of the methods (methods 1 and 4) resulted in the recovery of significantly less (P < 0.05) Salmonella when compared with the remaining 3 methods (methods 2, 3, and 5). No one method was successful in recovering Salmonella from all positive samples, although recovery with method 2 was statistically similar to the total number of positive samples analyzed (42 vs. 46 Salmonella-positive samples, P > 0.05). This study shows that culture methods differ significantly in their performance regarding the isolation of Salmonella from swine fecal samples.     

Isolation of Listeria monocytogenes from organs of young lambs

Samples of organs from 50 lambs, 3-45 days old, brought to this laboratory from 44 different flocks for post mortem examination, were examined for Listeria monocytogenes. Various diseases were found as cause of the death. The bacterium was isolated from 3 lambs (6%) which had died from E. coli-septicaemia. Two of these lambs were 3 days old whereas the third was 23 days.     

Isolation of Listeria monocytogenes from the skin of slaughtered beef cattle

We attempted to isolate Listeria monocytogenes from skin, contents of large intestines and carcasses of cattle introduced to a slaughterhouse in order to identify source of contamination for this pathogen. Sixty skin samples, 60 samples of the contents of large intestines and 30 carcass samples were colleted in June, August and November 2003 for use in this study. Listeria spp. and L. monocytogenes were isolated from 30 (50%) and 3 (5%) of the cattle skin samples, respectively. However, no Listeria spp., including L. monocytogenes, were isolated from intestinal contents or carcasses. Seven isolates were obtained, of which five and two strains were serotypes 1/2a and 1/2b, respectively. Genetic analysis suggested that there was persistent inhabitation of the pathogen around the area investigated in this study.     

食源性单增李斯特菌检测技术研究进展

单增李斯特菌是一种常见的食源性致病菌,会导致严重的人兽共患病。目前单增李斯特菌的检测方法主要包括传统方法、酶联免疫法、免疫胶体金技术、PCR检测技术、等温扩增技术、全自动微生物检测系统、生物芯片技术等。本文对各方法的原理、优缺点、应用情况及发展前景进行了概述。     

Listeria monocytogenes septicemia in a Thoroughbred foal.

Listeria monocytogenes septicemia was diagnosed in a 6-day-old Thoroughbred foal. Primary clinical signs included fever, depression, diarrhea, and respiratory distress. Hematologic abnormalities included leukopenia, neutropenia, degenerative left shift, and hyperfibrinogenemia. Clinical chemistry and blood gas abnormalities included metabolic acidosis, hypoxemia, hypocapnia, hypoglycemia, and hyponatremia. Despite aggressive therapeutic intervention and intensive care, the foal died within 12 hours of admission. A postmortem examination was performed, and the primary gross lesion was bilaterally severe, focally extensive bronchopneumonia. Histopathology revealed severe subacute multifocal suppurative bronchopneumonia with necrotizing vasculitis and intralesional coccobacilli. Cultures of blood collected at admission and immediately prior to death were positive for L. monocytogenes, as were cultures obtained from lung and liver at necropsy. Immunohistochemical examination of formalin-fixed tissues revealed abundant intra- and extracellular L. monocytogenes antigen within the lung and intravascularly in multiple organs.     

Studies on glutamic acid decarboxylase from Listeria monocytogenes.

The isolation and characterization of glutamic acid decarboxylase from Listeria monocytogenes has been described. Effects of various concentrations of glutamic acid as a substrate and pyridoxal phosphate as coenzyme on the activity of the partially purified enzyme have been examined and their Km and Vmax values determined. The enzyme exhibits relatively higher activity in 0.1 M pyridine-pyridine hydrochloride buffer with a pH value of 4.6.