High level of quinolone resistance in <Emphasis Type="Italic">Escherichia coli</Emphasis> from healthy chicken broilers

The development of resistance to quinolones (nalidixic acid, ciprofloxacin and enrofloxacin) in 2006–2008 was evaluated in 317 strains of Escherichia coli isolated from healthy chicken broilers from various farms. The isolates (2006/2007/2008) showed a high resistance to nalidixic acid (87/85/67 %), ciprofloxacin (CIP) (49/54/29 %) and enrofloxacin (ENR) (52/42/22 %). Nalidixic acid-resistant isolates with low level of MIC for CIP and ENR represented a single mutation; intermediary MIC for CIP and ENR were related to two mutations and high level resistance MIC for CIP (≥4 mg/L) and ENR (≥16 mg/L) represented three mutations (two in gyrA and one in parC). There was a correlation between the phenotype reading of high-level resistance and mutations in gyrA (Ser83Leu, Asp87Tyr or Asp87Asn) and parC (Ser80Ile) gene. Plasmid-mediated quinolone-resistance qnrS gene was detected in one Escherichia coli strain with a high level of ciprofloxacin resistance. Our results demonstrate the increase in occurrence of multiresistant E. coli strains with a high level of chromosomal and plasmid resistance to fluoroquinolones.     

Comparative studies for serodiagnosis of haemorrhagic septicaemia in cattle sera

Haemorrhagic septicaemia caused by Pasteurella multocida is a major epizootic disease in cattle and buffaloes in developing countries with high morbidity and mortality rate. In the present study, a total of 88 P. multocida isolates were isolated from 256 nasopharyngeal swabs and lung tissues samples (34.4%) during the period from January, 2013 to March, 2014 from different governorates located in Egypt. Dead calves showed the highest percentage of P. multocida isolation followed by the emergency slaughtered calves, diseased calves then apparently healthy ones. These isolates were confirmed as P. multocida microscopically, biochemically by traditional tests and by API 20E commercial kit then by PCR. The percentages of positive serum samples using somatic antigen and micro-agglutination test at 1/1280 diluted serum were 10%, 54.49% and 0% in apparently healthy, diseased and emergency slaughtered samples, respectively whereas, the percentages using capsular antigen and indirect haemagglutination test were 40%, 60.89% and 60% in apparently healthy, diseased and emergency slaughtered samples, respectively. The ELISA showed the highest sensitivity for diagnosing P. multocida in apparently healthy, diseased and emergency slaughtered animals with percentages of 42%; 92.9% and 80%, respectively. The obtained results revealed that the ELISA using capsular antigen of P. multocida is a more sensitive and specific serological test for diagnosis of haemorrhagic septicaemia.     

Adherence ofPasteurella multocida to porcine upper respiratory tract cells

A model to study the adherence ofPasteurella multocida to porcine upper respiratory tract cells is described. The ability of 27 differentP. multocida isolates to adhere to isolated tracheal epithelial cells was examined. The mean number of adherent bacterial cells was significantly greater (p<0.005) for capsular type A cells than for capsular type D cells. No significant differences were observed between toxigenic and nontoxigenic isolates, or between isolates exhibiting different somatic antigens. However, isolates from pigs without atrophic rhinitis showed only 65% of the adherence of isolates from pigs with atrophic rhinitis. Adherence ofP. multocida to porcine tracheal cells decreased with animal age; adherence to cells from adults was only half of the adherence to cells from newborn animals. The data indicate that, in the present experimental conditions, theP. multocida strains tested possess different abilities to attach to porcine upper respiratory tract cells.     

Lipase Activity from Strains of Pasteurella multocida

Thirteen clinical isolates of Pasteurella multocida from a variety of different animals and humans were examined for their ability to produce lipase. Lipase substrates used included Tween 20, Tween 40, Tween 80, and Tween 85. Lipase activity was detected in the filtrates of organisms grown to the exponential phase in Roswell Park Memorial Institute-1640 defined media (RPMI-1640), but activity increased in the filtrates when the cultures were allowed to proceed to the stationary phase. All strains examined (except for serotype 2) showed lipase activity against at least one of the Tweens. Tween 40 was the best substrate to demonstrate lipase activity. Pasteurella multocida serotype 8 produced the most active lipase against Tween 40 (3,561.7 units of activity/μg of protein). This activity continued to increase after P. multocida entered a stationary growth phase. P. multocida lipase activity was optimal at pH 8.0. Lipase activity of P. multocida serotype 8 was eluted from a Sepharose 2B column at several points, indicating that several lipases may be produced in vitro by this organism. These data demonstrate that clinical isolates of P. multocida produce lipase; therefore, this enzyme should be considered a potential virulence factors for this organism. Received: 16 September 1999 / Accepted: 22 November 1999     

牛源多杀性巴氏杆菌的分离与初步鉴定

【目的】本研究旨在对引起犊牛呼吸道综合征的多杀性巴氏杆菌进行分离鉴定,分析其亲缘关系和毒力基因的分布情况。【方法】收集2017年8月至2018年4月疑似患有犊牛呼吸道综合征的病牛鼻拭子进行细菌分离培养,对菌落形态和染色疑似巴氏杆菌的菌株进行16S rRNA测序和血清型鉴定,选择巴氏杆菌7类23种毒力基因,筛查临床分离株的毒力基因的分布。【结果】从8个省份的237份病料中分离出31株多杀性巴氏杆菌,分离率为13.1%。16S rRNA测序分析表明31株A型多杀性巴氏杆菌属于同一亚群,其序列同源性与中国分离株HB01以及国外分离株USDA-ARS-USMARC-60712、USDA-ARS-USMARC-60214、ATCC 43137以及36950亲缘关系较近。对分离出的31株A型多杀性巴氏杆菌的7类共23种毒力基因鉴定,结果显示31株多杀性巴氏杆菌所携带的毒力因子大多分布在17–19个,且集中度较高。【结论】A型多杀性巴氏杆菌为犊牛呼吸道综合症的主要流行血清型,通过对多杀性巴氏杆菌的临床分离株进化树和毒力基因分析,内蒙古、黑龙江、新疆、山西以及河北的7株分离株进化来源于同一分支,且均缺失毒力基因tadD和hgbA及携带毒力基因hsf-1,提示着其亲缘关系可能与其携带的特定毒力基因存在一定相关性。该研究为犊牛呼吸道综合征的病原学调查和多杀性巴氏杆菌流行病学调查提供了参考数据。     

Secretion of Proteases from Pasteurella multocida Isolates

The capability of Pasteurella multocida to secrete proteases to the culture medium and their characterization were studied in five animal isolates (bovine, chicken, sheep, and two from pig). All the isolates produced proteases in a wide range of molecular mass. It is suggested that they are neutral metalloproteases, since they were optimally active between pH 6 and 7, inhibited by chelating agents but not by other protease inhibitors, and reactivated by calcium. Proteases from isolates were able to degrade IgG. Several proteins from supernatants of cultures precipitated with 70% (NH₄)₂SO₄ of all the P. multocida isolates were recognized by a polyclonal antiserum raised against a purified protease from Actinobacillus pleuropneumoniae. Protease production might play an important role during tissue colonization and in P. multocida diseases. Received: 26 May 1998 / Accepted: 15 August 1998     

Serotyping and Enzyme Characterization of Pasteurella haemolytica and Pasteurella multocida Isolates Recovered from Pneumonic Lungs of Stressed Feeder Calves

Ninety-one isolates of Pasteurella multocida (Pm) and 124 of Pasteurella haemolytica (Ph) were recovered from the lungs of calves that died of bovine respiratory tract disease (BRTD). Nine Pm enzyme profiles (A through I) and 9 Ph enzyme profiles (J through R) were determined for the Pasteurella isolates. The Pm isolates were relatively evenly divided among the enzyme profiles, with one exception, profile I. The Ph isolates were not evenly distributed among the profiles. Fifty of the 91 Pm isolates were serotyped. Forty-two Pm isolates were positive for capsule type A, and 8 were untypable. Five somatic type antigen profiles (3; 3,4; 3,7; 3,4,7; and 4) were identified among the 50 serotyped Pm isolates; one isolate was untypable. The Ph isolates were further divided through serotyping and grouped as follows: 74 (60%) Pasteurella haemolytica A1 (PhA1), 12 (10%) PhA2, 4 (3%) PhA5, and 34 (27%) PhA6. Eighty-one percent of the Ph serotypes were clustered in the M and N enzyme profile. The P enzyme profile was almost unique to PhA2 (8 of 12, 67% of PhA2 isolates). Results of this study indicate a need to collect more data on Ph serotypes at the state veterinary diagnostic laboratories. Received: 29 August 1996 / Accepted: 15 October 1996     

Viral and Bacterial Pathogens in Bovine Respiratory Disease in Finland

Pathogens causing bovine respiratory tract disease in Finland were investigated. Eighteen cattle herds with bovine respiratory disease were included. Five diseased calves from each farm were chosen for closer examination and tracheobronchial lavage. Blood samples were taken from the calves at the time of the investigation and from 86 calves 3–4 weeks later. In addition, 6–10 blood samples from animals of different ages were collected from each herd, resulting in 169 samples. Serum samples were tested for antibodies to bovine parainfluenza virus-3 (PIV-3), bovine respiratory syncytial virus (BRSV), bovine coronavirus (BCV), bovine adenovirus-3 (BAV-3) and bovine adenovirus-7 (BAV-7). About one third of the samples were also tested for antibodies to bovine virus diarrhoea virus (BVDV) with negative results. Bacteria were cultured from lavage fluid and in vitro susceptibility to selected antimicrobials was tested. According to serological findings, PIV-3, BAV-7, BAV-3, BCV and BRSV are common pathogens in Finnish cattle with respiratory problems. A titre rise especially for BAV-7 and BAV-3, the dual growth of Mycoplasma dispar and Pasteurella multocida, were typical findings in diseased calves. Pasteurella sp. strains showed no resistance to tested antimicrobials. Mycoplasma bovis and Mannheimia haemolytica were not found.     

Identification of novel immunogens in <Emphasis Type="Italic">Pasteurella multocida</Emphasis>

   

P. multocida is a Gram-negative pathogen responsible for causing diseases in animals of economic significance to livestock industries throughout the world. Current vaccines include bacterins, which provide only limited protection against homologous serotypes. Therefore there is a need for more effective vaccines to control diseases caused by P. multocida. As a step towards developing vaccines against fowl cholera, a genomics based approach was applied for the identification of novel immunogens.     

In vitro potency and efficacy favor later generation fluoroquinolones for treatment of canine and feline Escherichia coli uropathogens in the United States

Information regarding in vitro activity of newer fluoroquinolones (FQs) is limited despite increasing resistance in canine or feline pathogenic Escherichia coli (E. coli). This study describes in vitro potency and efficacy toward E. coli of seven FQs grouped according to similarities in chemical structure: enrofloxacin, ciprofloxacin, orbifloxacin (first-group), levofloxacin, marbofloxacin (second-group) and pradofloxacin, moxifloxacin (third-group; latest S, S-pyrrolidino-piperidine at C-7). Potency measures included minimum inhibitory concentration (MIC) (geometric mean MIC, MIC₅₀, MIC₉₀); and mutant prevention concentration (MPC) for FQ susceptible isolates only. In vitro efficacy measures included relative susceptibility (MIC_BP-S:MIC) or resistance (MIC:MIC_BP-R) and mutant selection window (MSW) (MPC:MIC). For enrofloxacin susceptible isolates, mean MIC (μg/ml) was least for each third-group drug and ciprofloxacin and greatest for enrofloxacin and orbifloxacin (P = 0.006). For enrofloxacin susceptible isolates, MPC were below MIC:MIC_BP-R and least for pradofloxacin (0.29 ± 0.16 μg/ml) and greatest for enrofloxacin (1.55 ± 0.55 μg/ml) (P = 0.006). MSW was least for pradofloxacin (55 ± 30) and greatest for ciprofloxacin (152 ± 76) (P = 0.0024). MIC_BP-S:MIC was greatest (P = 0.025) for pradofloxacin (190.1 ± 0.61) and least for enrofloxacin (23.53 ± 0.83). For FQ susceptible isolates, FQs MIC:MIC_BP-R may serve as a surrogate for MPC. Because in vitro efficacy was greatest for pradofloxacin; it might be preferred for treatment of urinary tract infections (UTIs) associated with FQ susceptible E. coli uropathogens.     

Extracellular neuraminidase production by Pasteurella species isolated from infected animals

A total of 721 field isolates of various Pasteurella species (haemolytica, multocida, and testudinis) from various regions of the United States were examined for extracellular neuraminidase production. All strains were grown and tested in the same way. Included were 372 P. haemolytica serotype 1 isolates, 181 P. haemolytica serotype 2 isolates, 63 P. haemolytica serotype 6 isolates, 101 Pasteurella multocida isolates, and 4 Pasteurella testudinis isolates. All Pasteurella species examined produced the enzyme. The data revealed the following: (1) Several transfers of P. haemolytica strains on blood agar medium did not cause a decrease in enzyme activity. (2) P. haemolytica serotypes 2 and 6 produce more neuraminidase than P. haemolytica serotype 1, P. multocida, and P. testudinis isolates. (3) There was no apparent change in neuraminidase production by P. haemolytica serotypes 1 and 2 obtained from the same animal taken on different days in the feedyard. (4) There was no significant change in neuraminidase production by P. haemolytica serotype 2 isolates taken from the same animal at the auction market and later at the feedyard.     

Comparison of Active Drug Concentrations in the Pulmonary Epithelial Lining Fluid and Interstitial Fluid of Calves Injected with Enrofloxacin,Florfenicol, Ceftiofur,or Tulathromycin

Bacterial pneumonia is the most common reason for parenteral antimicrobial administration to beef cattle in the United States. Yet there is little information describing the antimicrobial concentrations at the site of action. The objective of this study was to compare the active drug concentrations in the pulmonary epithelial lining fluid and interstitial fluid of four antimicrobials commonly used in cattle. After injection, plasma, interstitial fluid, and pulmonary epithelial lining fluid concentrations and protein binding were measured to determine the plasma pharmacokinetics of each drug. A cross-over design with six calves per drug was used. Following sample collection and drug analysis, pharmacokinetic calculations were performed. For enrofloxacin and metabolite ciprofloxacin, the interstitial fluid concentration was 52% and 78% of the plasma concentration, while pulmonary fluid concentrations was 24% and 40% of the plasma concentration, respectively. The pulmonary concentrations (enrofloxacin + ciprofloxacin combined) exceeded the MIC₉₀ of 0.06 μg/mL at 48 hours after administration. For florfenicol, the interstitial fluid concentration was almost 98% of the plasma concentration, and the pulmonary concentrations were over 200% of the plasma concentrations, exceeding the breakpoint (≤ 2 μg/mL), and the MIC₉₀ for Mannheimia haemolytica (1.0 μg/mL) for the duration of the study. For ceftiofur, penetration to the interstitial fluid was only 5% of the plasma concentration. Pulmonary epithelial lining fluid concentration represented 40% of the plasma concentration. Airway concentrations exceeded the MIC breakpoint for susceptible respiratory pathogens (≤ 2 μg/mL) for a short time at 48 hours after administration. The plasma and interstitial fluid concentrations of tulathromcyin were lower than the concentrations in pulmonary fluid throughout the study. The bronchial concentrations were higher than the plasma or interstitial concentrations, with over 900% penetration to the airways. Despite high diffusion into the bronchi, the tulathromycin concentrations achieved were lower than the MIC of susceptible bacteria at most time points.     

Molecular characteristics and resistant mechanisms of imipenem-resistant <Emphasis Type="Italic">Acinetobacter baumannii</Emphasis> isolates in Shenyang,China

The investigation was carried out to elucidate the molecular characteristics and resistant mechanisms of imipenem-resistant Acinetobacter baumannii. Thirty-seven isolates were collected from January 2007 to December 2007. The homology of the isolates was analyzed by both pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The genes of β-lactamases, adeB, and class 1 integron were polymerase chain reaction amplified. Genotype analysis of the 37 A. baumannii isolates by PFGE revealed the circulation of four PFGE types (A-D); the A- and B-type accounted for 48.6% and 40.5%, respectively. MLST showed the existence of three allelic profiles. The agar dilution method was carried out to determine the MIC of imipenem, in the absence or presence of carbonyl cyanide m-chlorophenylhydrazone (CCCP, 10 μg/ml). The MICs of the strains to imipenem were between 16 μg/ml and 128 μg/ml. When CCCP was added, a MIC decrease of at least four-fold was observed in 20 isolates, which belonged to the A- or C-type. AdeB and bla _PER-1 genes were each detected in 35 isolates, bla _OXA-23 gene in 34 isolates and bla _OXA-58-like gene in 24 isolates. All isolates harbored bla _OXA-51-like genes. No isolates carried the bla _IMP-1 gene. Integron was detected in 25 isolates, which mediated the resistance to aminoglycosides and rifampin. The epidemiologic data suggested that the increasing infection of A. baumannii in our hospital was mainly caused by the inter-hospital spread of two epidemic clones. The AdeABC efflux system may be the important factor that leads to the high level of imipenem-resistance in PFGE A-type.     

In vivo production of neuraminidase by Pasteurella multocida A:3 in goats after transthoracic challenge

Six goats were injected transthoracically with live Pasteurella multocida A:3 to examine if an extracellular enzyme, neuraminidase, was produced in vivo during infection with this organism. The principal group of goats (n = 6) each received 1 ml of live 7.5 × 10⁴ cfu of P. multocida mixed with polyacrylate beads transthoracically in the left lung on day 0 and 1 ml of live P. multocida (2.2 × 10⁸ cfu) mixed with polyacrylate beads transthoracically in the left lung on day 22. Six goats were used as negative controls and received 0.3 g of polyacrylate beads subcutaneously in the right flank on days 0 and 22. Serum was obtained from all animals on days 0, 7, 14, 22, 29, and 36. Preimmune sera from all animals showed no detectable antibody to P. multocida A:3 neuraminidase in an enzyme neutralization assay. None of the sera from the negative control animals demonstrated a significant antibody titer against the P. multocida A:3 neuraminidase. On day 36, serum samples from the six infected animals possessed complete enzyme-neutralizing activity. Anti-neuraminidase antibody could be detected as early as day 14 in the infected animals. These data show that neuraminidase is produced in vivo during an active P. multocida A:3 lobar infection. Received: 16 March 1996 / Accepted: 19 April 1996     

Molecular characterization of the capsular antigens of Pasteurella multocida isolates using multiplex PCR

The use of molecular techniques for detection and characterization of the Pasteurella multocida is very important for rapid and specific detection and characterization of the organism. During the period from 15th February, 2014 to 15th April, 2015, 425 nasopharyngeal swabs and 175 lung and spleen samples were collected and examined by conventional methods, 80 strains (18.82%) of P. multocida were isolated from the calves, sheep and goat with respiratory manifestation. Meanwhile, 77 strains (44%) were isolated from emergency slaughtered animals. All the recovered strains were positive for specific PCR for detection of P. multocida strains previously identified as P. multocida by standard microbiological techniques. Multiplex PCR for molecular typing of the capsular antigens of the recovered P. multocida revealed positive amplification of 1044 bp fragments specific to the capsular antigen type A with 105 strains (66.88%), and amplification 511 bp fragments of the capsular antigen type E with 52 strain (33.12%) and absence of B, D and F antigens. Multiplex PCR for molecular typing of the capsular antigens of P. multocida can be used as a simple, sensitive, rapid, reliable technique instead of the serological techniques for identification of the capsular antigens of P. multocida     

Pasteurella multocida involved in respiratory disease of wild chimpanzees

Pasteurella multocida can cause a variety of diseases in various species of mammals and birds throughout the world but nothing is known about its importance for wild great apes. In this study we isolated P. multocida from wild living, habituated chimpanzees from Taï National Park, Côte d''Ivoire. Isolates originated from two chimpanzees that died during a respiratory disease outbreak in 2004 as well as from one individual that developed chronic air-sacculitis following this outbreak. Four isolates were subjected to a full phenotypic and molecular characterisation. Two different clones were identified using pulsed field gel electrophoresis. Multi Locus Sequence Typing (MLST) enabled the identification of previous unknown alleles and two new sequence types, ST68 and ST69, were assigned. Phylogenetic analysis of the superoxide dismutase (sodA) gene and concatenated sequences from seven MLST-housekeeping genes showed close clustering within known P. multocida isolated from various hosts and geographic locations. Due to the clinical relevance of the strains described here, these results make an important contribution to our knowledge of pathogens involved in lethal disease outbreaks among endangered great apes.     

Molecular diversity of porcine and human isolates of Pasteurella multocida

Aims: To examine the variability among Pasteurella multocida strains isolated from pigs (nasal, tonsil and lung specimens) and humans in France. Methods and Results: The genetic diversity of 117 French isolates of P. multocida, obtained from pigs (n = 101) and humans (n = 16) and three reference strains, was evaluated by pulsed‐field gel electrophoresis (PFGE) after macrorestriction with ApaI. Sixty‐four patterns were detected. The genetic relationships revealed five clusters (Aa1, Aa2, Aa3, Ab and B). The pig isolates obtained from pneumonic lungs and nasal cavities were clustered in groups Ab and Aa1, respectively (P < 0·05). Up to four different PFGE patterns were detected in the same farm. Isolates producing dermonecrotic toxins were clustered only in group Aa1, suggesting that the toxigenic isolates were more genetically homogenous than the others. Conversely, cluster Aa3 was significantly associated with human isolates even if the human isolates are spread over most of the clusters. Conclusions: Pasteurella multocida strains were genetically diverse, but pig and human isolates were significantly clustered in distinct phylogenetic groups. Significance and Impact of the Study: The discrimination index was >0·95 in both populations of human and pig isolates. Therefore, ApaI‐PFGE seems to be a useful tool for epidemiological tracing of P. multocida infections.     

基于响应面法的高抗原活性禽多杀性巴氏杆菌疫苗培养基的研制与效果评价

【背景】禽多杀性巴氏杆菌(Pasteurella multocida)引发的禽霍乱疫情造成了巨大的危害,而现有培养基存在培养菌密度较低的问题。【目的】研制高抗原活性的禽多杀性巴氏杆菌疫苗培养基。【方法】通过单因素试验、Plackett-Burman试验和响应面分析方法对禽多杀性巴氏杆菌培养基的成分进行调整,并对不同发酵阶段的菌体进行免疫原性测定。最后使用该培养基培养细菌后制备疫苗并通过动物攻毒试验评价其保护效果。【结果】使用研制的培养基培养禽多杀性巴氏杆菌,活菌密度能够在6 h达到约1.84×1010 CFU/mL,增菌效果是对照培养基的2.6倍;免疫原性测定结果显示在生长平台期菌体的抗原活性最高;攻毒试验表明制备的疫苗能够很好地抵抗禽多杀性巴氏杆菌的侵袭。【结论】研制出了高抗原活性的禽多杀性巴氏杆菌疫苗培养基,为疫苗的生产奠定了基础。     

Uterine Microbiology and Antimicrobial Susceptibility in Isolated Bacteria from Mares with Fertility Problems

Uterine microbiology and antimicrobial susceptibility was investigated in 239 mares with fertility problems in a prospective study in Sweden. Uterine swab samples were collected with double guarded swabs and transported overnight before being cultured. The Minimum Inhibitory Concentrations (MIC) was determined for a panel of antimicrobials. From 152 of the 239 mares at least one bacterial species was isolated, most frequently E. coli (104 isolates), β -haemolytic streptococci (31) and fungi (16). β -haemolytic streptococci were more frequently (p < 0.01) associated with clinical endometritis than with repeat breeding. The opposite was true for E. coli (p < 0.01). Among β -haemolytic streptococcal isolates some resistance was noted for 4 of 11 tested antibiotics, however, all isolates were susceptible to the widely used penicillin G. Among E. coli isolates enrofloxacin was the only of the 10 tested antibiotics for which no resistance was noted. Resistance was most commonly noted to cephalothin (39% of the isolates), streptomycin (22%), trimethoprim/sulphamethoxazole (15%) and ampicillin (11%). In conclusion, we show that both E. coli and β -haemolytic streptococci are frequently associated with fertility problems in mares and that antimicrobial resistance is a common feature of E. coli but also recognised for β -haemolytic streptococcal uterine isolates.     

Mercury Tolerance of Thermophilic <Emphasis Type="Italic">Bacillus</Emphasis> sp. and <Emphasis Type="Italic">Ureibacillus</Emphasis> sp

Although resistance of microorganisms to Hg(II) salts has been widely investigated and resistant strains have been reported from many eubacterial genera, there are few reports of mercuric ion resistance in extremophilic microorganisms. Moderately thermophilic mercury resistant bacteria were selected by growth at 62 °C on Luria agar containing HgCl₂. Sequence analysis of 16S rRNA genes of two isolates showed the closest matches to be with Bacillus pallidus and Ureibacillus thermosphaericus. Minimum inhibitory concentration (MIC) values for HgCl₂ were 80 μg/ml and 30 μg/ml for these isolates, respectively, compared to 10 μg/ml for B. pallidus H12 DSM3670, a mercury-sensitive control. The best-characterised mercury-resistant Bacillus strain, B. cereus RC607, had an MIC of 60 μg/ml. The new isolates had negligible mercuric reductase activity but removed Hg from the medium by the formation of a black precipitate, identified as HgS by X-ray powder diffraction analysis. No volatile H₂S was detected in the headspace of cultures in the absence or presence of Hg²⁺, and it is suggested that a new mechanism of Hg tolerance, based on the production of non-volatile thiol species, may have potential for decontamination of solutions containing Hg²⁺ without production of toxic volatile H₂S.