Subtilase Cytotoxin-Encoding subAB Operon Found Exclusively among Shiga Toxin-Producing Escherichia coli Strains

The presence of subAB was investigated for 3,453 Escherichia coli strains of various pathogenic categories. The occurrence of other virulence genes in subAB-positive strains was investigated. The subAB operon was detected among some Shiga toxin-producing E. coli (STEC) serotypes devoid of eae and carrying ehxA. Most subAB-positive strains also harbored stx₂, iha, saa, and lpfA_O113.Subtilase cytotoxin, a new member of the AB₅ toxin family, was identified for the first time in 2004 in a virulent O113:H21 Shiga toxin-producing Escherichia coli (STEC) strain that caused an outbreak of hemolytic-uremic syndrome in South Australia (16, 18). The presence of subAB genes was further detected in other STEC strains belonging to different serotypes (19). Subsequently, subAB genes were identified among STEC strains isolated in other countries (3, 8, 9, 14, 25).To evaluate how widely distributed the subAB operon is, we studied a large collection of STEC serotypes from nonhuman sources and E. coli strains of different pathogenic categories associated with human infections. The subAB-positive strains were further characterized regarding the presence of other virulence genes.A total of 2,255 E. coli strains isolated from humans and belonging to enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), extraintestinal pathogenic E. coli (ExPEC), and E. coli strains not belonging to the diarrheagenic categories described so far were randomly selected. STEC strains isolated in Brazil from humans have previously been tested for the presence of subAB by our group (3). The 1,198 STEC strains from nonhuman sources were isolated from dairy cattle, beef cattle, buffaloes, and goats. Overall, 109 different STEC serotypes were tested. An STEC strain of serotype O113:H21 (3) was used as a reference strain for subAB, cdt-V, and lpfA_O113, and E. coli strain DH5α was used as a negative control.The strains were screened for the presence of the subAB operon (encoding subtilase cytotoxin) using colony hybridization assays (21). The 1,823-bp subAB-specific DNA probe was derived from the STEC serotype O113:H21 (3) strain by PCR as previously described (19). Hybridization assays were performed under stringent conditions, and the probe was labeled with [α-³²P]dCTP (Amersham), using the Ready-To-Go DNA labeling kit (Amersham). All strains which yielded a positive or weak signal in hybridization assays with the subAB probe were retested by PCR (18, 19), and only those confirmed by PCR were considered to be carrying this sequence.The genetic profiles of the subAB-positive strains were determined using our previously reported data for the same strains (6, 7, 12, 13, 20, 24) regarding the presence of the ehxA, eae, stx₁, stx₂, and adhesin-encoding genes (1, 10, 11, 15, 17, 22, 23).A total of 130 STEC strains carrying the subAB operon, representative of each serotype and isolated from different animals, were analyzed by PCR for the presence of genes encoding Lpf_O113 and cytolethal distending toxin (Cdt-V) (2, 5).Expression of the SubAB and Cdt-V toxins was investigated using Chinese hamster ovary cells according to the methods of Paton et al. (18) and Bielaszewska et al. (2), respectively. Cells were exposed to filter-sterilized bacterial culture supernatants and observed daily for a period of 7 days. To confirm the loss of viability or morphological changes, trypan blue, violet crystal, and/or 4′,6-diamidino-2-phenylindole (DAPI) staining were performed. Control strains were included in all assays.As shown in Table ​Table1,1, the subAB operon was detected exclusively among STEC strains and corresponded to 25.5% (306/1,198) of the STEC collection. The presence of subAB was identified in 44.2% (141/319), 27.1% (29/107), and 23.8% (129/542) of STEC strains isolated from dairy cattle, buffaloes, and beef cattle, respectively. Only 3% (7/230) of STEC strains isolated from goat carried subAB. Among the 109 different STEC serotypes tested, 21 carried the subAB operon. The presence of the subAB operon probably is associated with some STEC serotypes. In the present study, the rate of carriage of this sequence within each serotype ranged from 5.5 to 100% (Table ​(Table2).2). Among caprine STEC strains, only those belonging to O113:H21 carried the subAB operon. A total of 306 (306/1,198) STEC strains carrying subAB were detected. We have previously reported that among 49 human STEC strains isolated in Brazil, none carried the subAB sequence (3).

TABLE 1.

E. coli strains of different pathogenic categories searched for the subAB operon

Celiac disease: its implications for orthopaedic nursing

Celiac disease (CD), also known as gluten-sensitive enteropathy, is an inherited autoimmune disease that can lead to multisystem effects. Left undiagnosed and untreated, it has severe orthopaedic implications involving osteoporosis and unnecessary treatment of joint pain, which could be avoided. The orthopaedic nurse may have thought that CD was primarily an abdominal disorder. We need to be aware of the potential practice implications in orthopaedics by understanding the disease, its diagnosis, and treatment. Furthermore, orthopaedic nurses can contribute to current research by studying the relationships between CD and orthopaedic problems.     

Diffusion-weighted magnetic resonance imaging findings in a patient with cerebral syphilitic gumma

Cerebral syphilitic gummas are rare entities, consisting of masses of granulation tissue that result from an exacerbated cell-mediated inflammatory response to Treponema pallidum, usually arising from the meninges of the convexity. Conventional magnetic resonance imaging and computed tomography findings of cerebral gummas have been reported, but diffusion-weighted imaging findings have not been previously described. In our patient, magnetic resonance imaging revealed a juxtacortical lesion with nodular enhancement, moderately restricted diffusion, a dural tail, and surrounding vasogenic edema. The role of diffusion-weighted imaging findings in the diagnosis of this condition is underscored.     

Boltushka: a homemade amphetamine-type stimulant and HIV risk in Odessa, Ukraine

BackgroundHomemade amphetamine-type stimulants (ATSs) have been reported in Russia and Eastern Europe for decades. Recipes differ geographically and over time producing differing active ingredients. Vint and jeff (active ingredients methamphetamine and methcathinone, respectively) are two such homemade ATSs originally produced from over-the-counter cold medications and household chemicals.MethodsDuring a Rapid Policy Assessment and Responses (RPAR) project in Odessa, Ukraine, researchers found use of boltushka, a novel homemade ATS. Fourteen supplemental qualitative interviews were conducted, including ten interviews with boltushka injectors and four interviews with pharmacists. We report patterns of boltushka use among local injection drug users (IDUs) as well as the role of laws, regulations, and current pharmacy practices.ResultsLegal restrictions on over-the-counter cold medicines in Ukraine led to products containing phenylpropanolamine (PPA), which oxidised with KMnO₄ (potassium permanganate), produces a weak ATS, cathinone, called boltushka. Boltushka’s ingredients are easily available in pharmacies or on the black market. IDUs reported a mean age at first use of 16 years old (range 12–21). While published data are scant, anecdotal evidence reported here include amphetamine-like effects on energy and appetite, binging patterns of use, and some reports of shaking and other neurological damage consistent with earlier reports from exposure to KMnO₄. Users reported sharing syringes and other non-sterile injection practices. No users reported specific treatment or prevention programs for boltushka users.ConclusionsAlthough Ukrainian government regulations have limited access to precursor chemicals, IDUs have continued to make and use boltushka. The actual extent and demographics of boltushka use are unknown. Besides risk of bloodborne disease, the health effects of injected homemade ATSs and their constituent chemicals are poorly documented. Interventions beyond available harm reduction efforts may be required. Education/treatment specific to boltushka users and screening for other physical harms are critical interventions.     

Drifting grating stimulation reveals particular activation properties of visual neurons in the caudate nucleus

The role of the caudate nucleus (CN) in motor control has been widely studied. Less attention has been paid to the dynamics of visual feedback in motor actions, which is a relevant function of the basal ganglia during the control of eye and body movements. We therefore set out to analyse the visual information processing of neurons in the feline CN. Extracellular single-unit recordings were performed in the CN, where the neuronal responses to drifting gratings of various spatial and temporal frequencies were recorded. The responses of the CN neurons were modulated by the temporal frequency of the grating. The CN units responded optimally to gratings of low spatial frequencies and exhibited low spatial resolution and fine spatial frequency tuning. By contrast, the CN neurons preferred high temporal frequencies, and exhibited high temporal resolution and fine temporal frequency tuning. The spatial and temporal visual properties of the CN neurons enable them to act as spatiotemporal filters. These properties are similar to those observed in certain feline extrageniculate visual structures, i.e. in the superior colliculus, the suprageniculate nucleus and the anterior ectosylvian cortex, but differ strongly from those of the primary visual cortex and the lateral geniculate nucleus. Accordingly, our results suggest a functional relationship of the CN to the extrageniculate tecto-thalamo-cortical system. This system of the mammalian brain may be involved in motion detection, especially in velocity analysis of moving objects, facilitating the detection of changes during the animal's movement.