The description of an esculin-positive biovar of Pseudomonas aeruginosa

In the study of 280 P. aeruginosa strains isolated in different hospitals of St. Petersburg for the first time 48 strains capable of hydrolyzing esculin have been detected. The hydrolysis of esculin is determined in plates with the use of the microvolume techniques the results were evaluated after 3-hour incubation at 37 degrees C. The data confirming the existence of the exculin-positive biovar of P. aeruginosa have been obtained; these data show the wide spread of esculin-positive strains in hospitals of different specialization (17.1 +/- 5.1% of P. aeruginosa strains), the characteristic combination of the sign of esculin hydrolysis with such signs as the absence of the smell of trimethylamine and the phenomenon of "iridescent lysis" of the colonies, the stability of the sign of esculin hydrolysis in strains, repeatedly isolated from patients, after the storage of the cultures and their treatment with plasmid-eliminating preparation. The name "esculinolytica" has been proposed for this biovar. The typing strain of biovar esculinolytica has been deposited in the culture collection of the Russian Research Institute of Agricultural Microbiology as P. aeruginosa ARRIAM 64-A. This biovar been found to be most widely spread in urological hospitals, where esculin-positive strains are isolated 3 times more frequently (32.2 +/- 5.1% of P. aeruginosa strains) than in surgical hospitals (10.7 +/- 2.2%).     

Isolation and characterization of a lipolytic bacterium capable of growing in a low-water-content oil-water emulsion.

A unique lipolytic bacterium was isolated in a selective growth system consisting of 99% triglycerides and a 1% water phase. The bacterium, termed Pseudomonas aeruginosa YS-7, was able to grow in an environment of low water content and could also survive amphipathic, osmotic, and matrical water stress in a triglyceride-rich culture. The isolated strain was identified as P. aeruginosa on the basis of standard physiological, biochemical, and serological assays. The strain is a gram-negative motile rod, aerobic, pigment forming, and capable of growing at 42 degrees C. It is highly tolerant of high concentrations of the cationic detergent cetyltrimethylammonium bromide and of the fatty acid salts derived from bacterial hydrolysis of the oil. Growth of the bacterium in a pure culture in a 99% triglyceride medium lasted until most of the water was evaporated or consumed. Growth was accompanied by triglyceride hydrolysis, which continued to occur even after growth saturation until the water was totally depleted. No loss of viability was observed when the culture was maintained under water-depleted conditions for an additional 40 h. A second cycle of bacterial growth and triglyceride hydrolysis was immediately initiated upon the addition of 1% (vol/vol) water to the culture. Lipase activity was stable regardless of changes in culture conditions. The isolated strain is uniquely resistant to severe water stress in a triglyceride-rich medium or under cold acetone precipitation compared with 12 other microbial strains, including bacteria and yeasts. Among these 12, only the lipolytic strains grew in the 99% triglyceride medium, but they reached a cell mass fourfold smaller than that of P. aeruginosa YS-7.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation and characterization of a lipolytic bacterium capable of growing in a low-water-content oil-water emulsion

A unique lipolytic bacterium was isolated in a selective growth system consisting of 99% triglycerides and a 1% water phase. The bacterium, termed Pseudomonas aeruginosa YS-7, was able to grow in an environment of low water content and could also survive amphipathic, osmotic, and matrical water stress in a triglyceride-rich culture. The isolated strain was identified as P. aeruginosa on the basis of standard physiological, biochemical, and serological assays. The strain is a gram-negative motile rod, aerobic, pigment forming, and capable of growing at 42 degrees C. It is highly tolerant of high concentrations of the cationic detergent cetyltrimethylammonium bromide and of the fatty acid salts derived from bacterial hydrolysis of the oil. Growth of the bacterium in a pure culture in a 99% triglyceride medium lasted until most of the water was evaporated or consumed. Growth was accompanied by triglyceride hydrolysis, which continued to occur even after growth saturation until the water was totally depleted. No loss of viability was observed when the culture was maintained under water-depleted conditions for an additional 40 h. A second cycle of bacterial growth and triglyceride hydrolysis was immediately initiated upon the addition of 1% (vol/vol) water to the culture. Lipase activity was stable regardless of changes in culture conditions. The isolated strain is uniquely resistant to severe water stress in a triglyceride-rich medium or under cold acetone precipitation compared with 12 other microbial strains, including bacteria and yeasts. Among these 12, only the lipolytic strains grew in the 99% triglyceride medium, but they reached a cell mass fourfold smaller than that of P. aeruginosa YS-7.(ABSTRACT TRUNCATED AT 250 WORDS)     

Detection of lipase in Vibrio cholerae serogroup O1 in reaction of volumetric agglomeration

Study showed that El-Tor strains of V. cholerae isolated from different sources produce lipase for hemolysis after cultivation during 24 h on meat-peptone broth independently from their toxigenic and hemolytic abilities. Study of 3- and 4-hours broth cultures of vibrios revealed possibility to differentiate between hemolytic nontoxigenic strains and toxigenic nonhemolytic ones. Using antilipaze diagnostic kit it was possible to differentiate El-Tor vibrios from vibrios of classic biovar basing on lipase production 24 h after cultivation on meat-peptone broth that was evident in El-Tor vibrios but not in classic biovar strains.     

Drug susceptibility of Pseudomonas aeruginosa strains isolated from patients of a hospital and specialistic outpatients clinics of the SP ZOZ in Nidzica

The aim of this study was to evaluate a frequency of isolation and antimicrobial susceptibility testing (AST) of Pseudomonas aeruginosa strains cultured from clinical specimens collected from patients hospitalized in wards and specialistic outpatients clinics of a hospital in Nidzica (01. 09. 2000 -31. 12. 2003). During over three years 392 Pseudomonas aeruginosa strains were cultured from 16346 clinical samples provided to bacteriological laboratory. P. aeruginosa strains were isolated from 2.5% of examined specimens. Susceptibility of Pseudomonas aeruginosa strains to antimicrobial agents was tested. The highest in vitro activity against clinical P. aeruginosa strains demonstrated imipenem. One strain was resistant to imipenem. This strain was isolated from a patient of a surgical department. Metalo-beta-lactamase was not detected (MBL-negative strain).Twenty nine strains were ESBL producer (7.4% of all strains). The contribution of Pseudomonas aeruginosa strains to the etiology of nosoconial and ambulatory infections increases. In vitro activity of antibacterial agents against P. aeruginosa strains should be monitored during therapy of infections. Resistance to antibiotics/chemothe-rapeutics may be acquired during treatment with antibacterial agent to which P. aeruginosa strain was susceptible according to the antibiogram.     

Occurrence and susceptibility to antibiotics of carbapenem-resistant Pseudomonas aeruginosa strains between 1998 and 2009

P. aeruginosa rods are dangerous pathogens mainly responsible for nosocomial infections of different localization. Resistance to carbapenems, observed among them, is a serious threat due to ability to be transmitted between bacterial species. The aim of our study was to retrospectively evaluate the frequency of isolation and susceptibility to antibiotics of imipenem- and meropenem-resistant P. aeruginosa strains isolated between 1998 and 2009 from patients of University Hospital No 1 of dr A. Jurasz in Bydgoszcz. Study shows increasing number of isolation that type of strains from 19 in 1998 to 144 in 2009. Among all isolated P. aeruginosa strains majority was obtained from patients of the Intensive Care Units, Rehabilitation and Surgery Clinics. Examined strains of P. aeruginosa rods were mainly isolated from urine (20.5%), bronchoalveolar lavage (17.7%) and wound swabs (14.5%) samples. The isolates demonstrated frequently resistance to carbenicillin (> or 66.7%), ticarcillin (> or = 72.7%) and cefotaxime (> or = 75.6%). The lowest rate of resistant strains was observed in case of ceftazidime (< or = 68.8%), aztreonam (< or = 47.4%) and colistin (< or = 1.7%) suggesting the highest activity of that antimicrobials against infections caused by examined strains.     

A study of the incidence of different fluorescent Pseudomonas species and biovars in the microflora of fresh and spoiled meat and fish, raw milk, cheese, soil and water.

Of 182 various foodstuffs and environmental samples examined, 86% had microflora containing fluorescent Pseudomonas in differing proportions. A computer-aided technique was used to identify most of the 445 fluorescent strains. Pseudomonas fluorescens biovar V-1 was most frequently isolated (24%); it either predominated or was present in all types of samples. Other strains, belonging to the other subgroups of biovar V (V-2, V-4, V-5, V-6 and V-7), together represented 14.3%. We also identified Ps. fluorescens biovars I-1 and I-2 (13.9%), II-1 and II-3 (3.6%), III-1 and III-2 (8.7%), IV-2 (0.7%); Ps. putida A and B (11%); Ps. lundensis (10.3%); group B3 (2%) and Ps. aeruginosa (0.7%). Unidentified strains accounted for 10.6% of the flora, many resembling Ps. fluorescens biovar V. Although the presence of Ps. fluorescens V-1 was often marked, other taxa predominated or were present in large quantities in some particular samples, such as Ps. fluorescens I-1 in raw milk and cheese, Ps. lundensis in spoiled meat and Ps. fluorescens III-1 in spoiled fish. Pseudomonas putida A and B were evident in environmental rather than in food samples.     

Comparison of flagellin genes from clinical and environmental Pseudomonas aeruginosa isolates

Pseudomonas aeruginosa, an important opportunistic pathogen, was isolated from environmental samples and compared to clinically derived strains. While P. aeruginosa was isolated readily from an experimental mushroom-growing unit, it was found only rarely in other environmental samples. A flagellin gene PCR-restriction fragment length polymorphism analysis of the isolates revealed that environmental and clinical P. aeruginosa strains are not readily distinguishable. The variation in the central regions of the flagellin genes of seven of the isolates was investigated further. The strains used included two strains with type a genes (998 bp), four strains with type b genes (1,258 bp), and one strain, K979, with a novel flagellin gene (2,199 bp). The route by which flagellin gene variation has occurred in P. aeruginosa is discussed.     

A study of the incidence of different fluorescent Pseudomonas species and biovars in the microflora of fresh and spoiled meat and fish, raw milk, cheese, soil and water

M. GENNARI AND F. DRAGOTTO. 1992. Of 182 various foodstuffs and environmental samples examined, 86% had microflora containing fluorescent Pseudomonas in differing proportions. A computer-aided technique was used to identify most of the 445 fluorescent strains. Pseudomonas fluorescens biovar V-1 was most frequently isolated (24%); it either predominated or was present in all types of samples. Other strains, belonging to the other subgroups of biovar V (V-2, V-4, V-5, V-6 and V-7), together represented 14.3%. We also identified Ps. fluorescens biovars I-1 and I-2 (13.9%), II-1 and II-3 (3.6%), III-1 and III-2 (8.7%), IV-2 (0.7%); Ps. putida A and B (11%); Ps. lundensis (10.3%); group B3 (2%) and Ps. aeruginosa (0.7%). Unidentified strains accounted for 10.6% of the flora, many resembling Ps. fluorescens biovar V. Although the presence of Ps. fluorescens V-1 was often marked, other taxa predominated or were present in large quantities in some particular samples, such as Ps. fluorescens I-1 in raw milk and cheese, Ps. lundensis in spoiled meat and Ps. fluorescens III-1 in spoiled fish. Pseudomonas putida A and B were evident in environmental rather than in food samples.     

Carbenicillin-hydrolyzing penicillinases of Proteus mirabilis and the PSE-type penicillinases of Pseudomonas aeruginosa

Three carbenicillin-hydrolyzing penicillinases were found in Proteus mirabilis strains, N-3, N-29, and GN79. The former two strains were isolated in 1978, but strain GN79 was one of our stock cultures isolated in 1965. These penicillinases closely resembled each other, and the PSE-1 and PSE-4 enzymes produced by Pseudomonas aeruginosa, in their substrate profiles and kinetic properties for hydrolyzing various beta-lactams. However, differences were found in their molecular weights and isoelectric points which ranged from 22,000 to 27,000 and from 6.0 to 6.9, respectively. The antiserum against the purified penicillinase of N-29 cross-reacted with the enzyme of N-3 and inhibited its activity by more than 80%. The antiserum also reacted with the PSE-1 and PSE-4 enzymes. The antiserum did not react with the penicillinase from strain GN79 and the PSE-2 and PSE-3 enzymes of P. aeruginosa. Enzyme production in N-3 and N-29 was mediated by R plasmids.     

Species of Pseudomonas obtained at 7 degrees C and 30 degrees C during aerobic storage of lamb carcasses.

A total of 268 strains of Pseudomonas isolated during storage life of lamb carcasses was identified to species level. One-hundred and thirteen strains obtained at 30 degrees C were Ps. fragi (51), Ps. lundensis (17), Ps. fluorescens biovars I (10), III (9) and VI (1), Ps. putida biovar A (8 strains) and unidentified (17 strains). Species and biovars isolated at 7 degrees C (155) were Ps. fragi (101), Ps. lundensis (32), Ps. fluorescens biovar I (6), Ps. putida biovar A (8) and unidentified (8). Numerical analysis (82% SSM, UPGMA) of 'psychrotrophic' and 'mesophilic' strains resulted in the formation of nine and eight clusters respectively. The dendrograms obtained exhibited similar structures. Most of the strains of Ps. lundensis and Ps. fragi clustered together. Strains of this latter species also joined the type strain of Ps. testosteroni and appeared included with phenons containing the Ps. putida strains. There were clusters made up exclusively of strains assigned to one biovar or group (Ps. fluorescens biovars I and II and unidentified). A high level of similarity was observed between clusters of Ps. fluorescens biovar I and those containing the Ps. fragi-Ps. lundensis complex (> 74% SSM) and Ps. lundensis (> 80%). The recovery of pseudomonads seemed to be affected by the sampling day.     

Species of Pseudomonas obtained at 7°C and 30°C during aerobic storage of lamb carcasses

A total of 268 strains of Pseudomonas isolated during storage life of lamb carcasses was identified to species level. One-hundred and thirteen strains obtained at 30°C were Ps.fragi (51), Ps. lundensis (17), Ps. fluorescens biovars I (10), III (9) and VI (1), Ps. putida biovar A (8 strains) and unidentified (17 strains). Species and biovars isolated at 7°C (155) were Ps. fragi (101), Ps. lundensis (32), Ps. fluorescens biovar I (6), Ps. putida biovar A (8) and unidentified (8). Numerical analysis (82% S _SM, UPGMA) of 'psychrotrophic' and 'mesophilic' strains resulted in the formation of nine and eight clusters respectively. The dendrograms obtained exhibited similar structures. Most of the strains of Ps. lundensis and Ps. fragi clustered together. Strains of this latter species also joined the type strain of Ps. testosteroni and appeared included with phenons containing the Ps. putida strains. There were clusters made up exclusively of strains assigned to one biovar or group ( Ps. fluorescens biovars I and II and unidentified). A high level of similarity was observed between clusters of Ps. fluorescens biovar I and those containing the Ps. fragi-Ps. lundensis complex (>74% S _SM) and Ps. lundensis (>80%). The recovery of pseudomonads seemed to be affected by the sampling day.     

TOL plasmid in Pseudomonas aeruginosa PAO: thermosensitivity of self-maintenance and inhibition of host cell growth.

The TOL plasmid originally isolated in Pseudomonas putida (arvilla) mt-2 was transmissible to strains of the fluorescens group of Pseudomonas, i.e., P. putida, P. fluorescens, and P. aeruginosa, except for a strain of P. aeruginosa, strain PAO. The same strain, however, could accept the plasmid when its restriction and modification abilities were lost by mutations or by growing at high temperature. In addition, the transmissibility of the TOL plasmid from strain PAO to P. putida was low when the plasmid was modified by the donor. By using P. aeruginosa PAO carrying the TOL plasmid, the stability and genetic expression of the plasmid as well as its effect on the host cell growth were examined. Thus the self-maintenance of the plasmid was found to be thermosensitive. Furthermore, the TOL plasmid inhibited the growth of strain PAO at high temperature, accompanied by the formation of some filamentous cells. These thermosensitive properties of the TOL plasmid were host dependent and not exhibited in another strain of P. aeruginosa.     

Selective medium for Pseudomonas aeruginosa that uses 1,10-phenanthroline as the selective agent.

The MIC of 1,10-phenanthroline for 35 Pseudomonas aeruginosa strains was 128 micrograms/ml, whereas 32 micrograms or less per ml inhibited all other microorganisms tested. On the basis of these results, a selective agar for P. aeruginosa which contained 15 g of Trypticase soy broth (BBL Microbiology Systems), 15 g of agar, and 0.1 g of phenanthroline per liter was formulated. Forty-four P. aeruginosa strains yielded a mean efficiency of plating on this medium of 79% of the counts obtained on Trypticase soy agar, which was significantly higher than that obtained with pseudomonas isolation agar or Pseudosel agar. Pseudomonas cepacia, Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas stutzeri, representatives of 13 other genera (including gram-negative rods, gram-positive rods, and cocci), and a yeast were not recovered within 48 h at 35 degrees C when approximately 10(7) CFU were plated on this medium. Only small colonies from one strain each of P. fluorescens and P. putida could be seen at 3 and 7 days, respectively, and they had an efficiency of plating of only less than 0.001%. When 10(7) CFU of either of these strains was plated with 10(2) CFU of P. aeruginosa, it did not interfere with the quantitative recovery of P. aeruginosa.     

Selective medium for Pseudomonas aeruginosa that uses 1,10-phenanthroline as the selective agent

The MIC of 1,10-phenanthroline for 35 Pseudomonas aeruginosa strains was 128 micrograms/ml, whereas 32 micrograms or less per ml inhibited all other microorganisms tested. On the basis of these results, a selective agar for P. aeruginosa which contained 15 g of Trypticase soy broth (BBL Microbiology Systems), 15 g of agar, and 0.1 g of phenanthroline per liter was formulated. Forty-four P. aeruginosa strains yielded a mean efficiency of plating on this medium of 79% of the counts obtained on Trypticase soy agar, which was significantly higher than that obtained with pseudomonas isolation agar or Pseudosel agar. Pseudomonas cepacia, Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas stutzeri, representatives of 13 other genera (including gram-negative rods, gram-positive rods, and cocci), and a yeast were not recovered within 48 h at 35 degrees C when approximately 10(7) CFU were plated on this medium. Only small colonies from one strain each of P. fluorescens and P. putida could be seen at 3 and 7 days, respectively, and they had an efficiency of plating of only less than 0.001%. When 10(7) CFU of either of these strains was plated with 10(2) CFU of P. aeruginosa, it did not interfere with the quantitative recovery of P. aeruginosa.     

Numerical taxonomy of fluorescent Pseudomonas associated with tomato roots

The phenetic taxonomy of 110 fluorescent bacterial strains, isolated from the roots of tomatoes and other plants was numerically studied through 97 features including 69 assimilation tests. Thirty-two reference strains of various Pseudomonas spp. were additionally included. The strains clustered into 16 clusters at the 74% similarity level when using Jaccard similarity coefficients. Almost all field strains belonged to the P. fluorescens/P. putida-complex while none clustered with P. syringae and allied bacteria. The biovar II branch, as well as the newly described biovar VI of P. fluorescens, made up 55% and 20% respectively, of the field strains; two % were allocated to P. fluorescens biovar I and three % to biovar IV. Eleven % of the root associated strains were designated P. putida; six strains were biovar A, three strains biovar B while four strains could not be referred to any known biovar. The continuum within the P. fluorescens/P. putida-complex as well as the taxonomic status of the six biovars of P. fluorescens and the three biovars of P. putida are discussed.     

Recognition of biovar C of Fusobacterium necrophorum (Flügge) Moore and Holdeman as Fusobacterium pseudonecrophorum sp. nov., nom. rev. (ex Prévot 1940)

The cellular morphology, colonial morphology, biochemical properties, DNA base compositions, and DNA-DNA homolgies of three biovars of Fusobacterium necrophorum were examined. Some differences were found among the three biovars in cellular morphology, colonial morphology, and biochemical properties. The guanine-plus-cytosine contents of DNAs from biovar C strains Fn521T (T = type strain), Fn522, and Fn520 were 30.4, 29.3, and 28.0 mol%, respectively, and the guanine-plus-cytosine contents of DNAs from strains VPI 2891 (biovar A) and VPI 6161 (biovar B) were 31.3 and 32.0 mol%, respectively. Labeled DNA from biovar C strain Fn521T exhibited 96 and 82% relatedness to DNAs from biovar C strains Fn522 and Fn520, respectively; however, it exhibited only about 10% relatedness to DNAs from strains of biovars A and B. Labeled DNAs from strains VPI 2891 and VPI 6161 exhibited more than 70% relatedness to each other, but about 6 to 20% relatedness to DNAs from biovar C strains. Therefore, Fusobacterium pseudonecrophorum sp. nov., nom. rev. (ex Prévot 1940) is proposed for Fusobacterium necrophorum biovar C. The type strain is strain Fn521 (= JCM 3722).     

铜绿假单胞菌对碳青霉烯类药物的耐药机制研究

目的 探讨铜绿假单胞菌对碳青霉烯类药物的耐药机制.方法 收集2008年11月至2009年4月我院临床分离的铜绿假单胞菌31株,根据药敏结果分为碳青霉烯类耐药组(21株)和碳青霉烯类敏感组(10株).另设1株标准株ATCC 27853,用亚胺培南-EDTA(乙二胺四乙酸)抑制试验检测菌株是否产生金屑酶,采用PCR法检测各菌株的外膜孔道蛋白oprD2基因,探讨铜绿假单胞菌对碳青霉烯类抗生素耐药机制.结果 21株耐药株有7株产生金属酶;21株耐药株经oprD2基因扩增,15株阴性,6株阳性,10株敏感株全部阳性.统计学检验结果表明,碳青霉烯类耐药组与敏感组oprD2基因阳性率的差异有极显著性(P＜0.01).结论 oprD2基因缺失和金属酶是本院铜绿假单胞菌对碳青霉烯类抗生素耐药的重要机制.     

Fatty acid composition of lipid A in Pseudomonas fluorescens

The fatty acid composition of lipid A was studied using gas-liquid chromatography (GLC) and GLC-mass spectrometry in Pseudomonas fluorescens strains of biovars A, B, C, i, F and G, the type strain ATCC 13525 (biovar A) inclusive. The following fatty acids were identified as predominant in the composition of lipid A in the strains representing biovars A, B, C, i, F and G: 3-hydroxydecanoic (3-OH C10:0), 2-hydroxydodecanoic (2-OH C12:0), 3-hydroxydodecanoic (3-OH C12:0), dodecanoic (C12:0), hexadecanoic (C16:0), octadecanoic (C18:0), hexadecenoic (C16:1) and octadecenoic (C18:1) acids. Lipid A of a biovar G strain differed noticeably from other strains in its fatty acid composition. Its main components were as follows: 3-hydroxytetradecanoic (3-OH C14:0), 3-hydroxypentadecanoic (3-OH C15:0) and dodecanoic (C12:0) fatty acids. The coefficients of similarity were determined for lipid A specimens isolated from the studied strains of P. fluorescens by calculating their fatty acid composition with a computer.     

Isolation of Vibrio alginolyticus and Vibrio splendidus from aquacultured carpet shell clam (Ruditapes decussatus) larvae associated with mass mortalities

Two episodes of mortality of cultured carpet shell clams (Ruditapes decussatus) associated with bacterial infections were recorded during 2001 and 2002 in a commercial hatchery located in Spain. Vibrio alginolyticus was isolated as the primary organism from moribund clam larvae that were obtained during the two separate events. Vibrio splendidus biovar II, in addition to V. alginolyticus, was isolated as a result of a mixed Vibrio infection from moribund clam larvae obtained from the second mortality event. The larval mortality rates for these events were 62 and 73%, respectively. Mortality was also detected in spat. To our knowledge, this is the fist time that these bacterial species have been associated with larval and juvenile carpet shell clam mortality. The bacterial strains were identified by morphological and biochemical techniques and also by PCR and sequencing of a conserved region of the 16S rRNA gene. In both cases bacteria isolated in pure culture were inoculated into spat of carpet shell clams by intravalvar injection and by immersion. The mortality was attributed to the inoculated strains, since the bacteria were obtained in pure culture from the soft tissues of experimentally infected clams. V. alginolyticus TA15 and V. splendidus biovar II strain TA2 caused similar histological lesions that affected mainly the mantle, the velum, and the connective tissue of infected organisms. The general enzymatic activity of both live cells and extracellular products (ECPs), as evaluated by the API ZYM system, revealed that whole bacterial cells showed greater enzymatic activity than ECPs and that the activity of most enzymes ceased after heat treatment (100 degrees C for 10 min). Both strain TA15 and strain TA2 produced hydroxamate siderophores, although the activity was greater in strain TA15. ECPs from both bacterial species at high concentrations, as well as viable bacteria, caused significant reductions in hemocyte survival after 4 h of incubation, whereas no significant differences in viability were observed during incubation with heat-killed bacteria.