Development of an efficient entire-capsid-coding-region amplification method for direct detection of poliovirus from stool extracts.

Laboratory diagnosis has played a critical role in the Global Polio Eradication Initiative since 1988, by isolating and identifying poliovirus (PV) from stool specimens by using cell culture as a highly sensitive system to detect PV. In the present study, we aimed to develop a molecular method to detect PV directly from stool extracts, with a high efficiency comparable to that of cell culture. We developed a method to efficiently amplify the entire capsid coding region of human enteroviruses (EVs) including PV. cDNAs of the entire capsid coding region (3.9 kb) were obtained from as few as 50 copies of PV genomes. PV was detected from the cDNAs with an improved PV-specific real-time reverse transcription-PCR system and nucleotide sequence analysis of the VP1 coding region. For assay validation, we analyzed 84 stool extracts that were positive for PV in cell culture and detected PV genomes from 100% of the extracts (84/84 samples) with this method in combination with a PV-specific extraction method. PV could be detected in 2/4 stool extract samples that were negative for PV in cell culture. In PV-positive samples, EV species C viruses were also detected with high frequency (27% [23/86 samples]). This method would be useful for direct detection of PV from stool extracts without using cell culture.

Multiple independent emergences of type 2 vaccine-derived polioviruses during a large outbreak in northern Nigeria.

Since 2005, a large poliomyelitis outbreak associated with type 2 circulating vaccine-derived poliovirus (cVDPV2) has occurred in northern Nigeria, where immunization coverage with trivalent oral poliovirus vaccine (tOPV) has been low. Phylogenetic analysis of P1/capsid region sequences of isolates from each of the 403 cases reported in 2005 to 2011 resolved the outbreak into 23 independent type 2 vaccine-derived poliovirus (VDPV2) emergences, at least 7 of which established circulating lineage groups. Virus from one emergence (lineage group 2005-8; 361 isolates) was estimated to have circulated for over 6 years. The population of the major cVDPV2 lineage group expanded rapidly in early 2009, fell sharply after two tOPV rounds in mid-2009, and gradually expanded again through 2011. The two major determinants of attenuation of the Sabin 2 oral poliovirus vaccine strain (A481 in the 5'-untranslated region [5'-UTR] and VP1-Ile143) had been replaced in all VDPV2 isolates; most A481 5'-UTR replacements occurred by recombination with other enteroviruses. cVDPV2 isolates representing different lineage groups had biological properties indistinguishable from those of wild polioviruses, including efficient growth in neuron-derived HEK293 cells, the capacity to cause paralytic disease in both humans and PVR-Tg21 transgenic mice, loss of the temperature-sensitive phenotype, and the capacity for sustained person-to-person transmission. We estimate from the poliomyelitis case count and the paralytic case-to-infection ratio for type 2 wild poliovirus infections that ∼700,000 cVDPV2 infections have occurred during the outbreak. The detection of multiple concurrent cVDPV2 outbreaks in northern Nigeria highlights the risks of cVDPV emergence accompanying tOPV use at low rates of coverage in developing countries.

Arrhythmogenicity of hypothermia - a large animal model of hypothermia.

BACKGROUND: Therapeutic hypothermia (TH) is used to mitigate cerebral injury after an out of hospital cardiac arrest. There is a perceived risk of increased arrhythmias with temperatures lower than the current target of 32-34°C for TH. This study sought to develop and investigate the electrophysiological changes in a sheep model of systemic hypothermia regarding the susceptibility to ventricular arrhythmias. METHODS: Ten sheep underwent systemic hypothermia using a venous-venous extra-corporeal circuit whilst instrumented with a 12 lead ECG. An epicardial sock recorded potentials to 30°C (N=10) or 26°C (N=6). Activation times (AT) and Activation Recovery Intervals (ARI) were calculated using custom software. RESULTS: The AT and ARI were significantly prolonged with increased heterogeneity during hypothermia. This effect was most pronounced between normothermia and 34°C during sinus rhythm (SR). For ventricular pacing (VP) however heterogeneity continued to increase with progressive hypothermia. CONCLUSIONS: Hypothermia causes a significant increase in the heterogeneity of depolarisation and repolarisation. There is evidence to suggest that SR is protective with most of the increase in heterogeneity occurring with cooling to 34°C. This raises the possibility that the current target temperatures for therapeutic hypothermia may be safely lowered to provide a gain in cerebral protection.

Ficolin-2 and ficolin-3 in women with malignant and benign ovarian tumours.

Ficolins are serum pattern recognition molecules. They have opsonic properties and are able to activate complement via the lectin pathway. This paper reports investigations concerning ficolin-2 and ficolin-3 in ovarian cancer (OC). Their serum levels, single nucleotide polymorphisms of the corresponding FCN2 and FCN3 genes and specific mRNA expression in ovarian sections were investigated in 128 patients suffering from primary OC and 197 controls operated on for reasons other than malignancies. The latter consisted of two reference groups: those with benign tumours (n = 123) and those with normal ovaries (NO) (n = 74). Serum ficolin-2 and ficolin-3 concentrations were higher among patients with malignant disease when compared with either of the reference groups. A significant correlation between ficolin-2 and ficolin-3 concentrations was found, while no correlations with CA125 antigen or CRP were observed. No differences in the frequency of single nucleotide polymorphisms at sites -64, -4 (promoter), +6359, or +6424 (exon 8) (FCN2 gene) nor in the frame-shift mutation 1637delC (FCN3 gene) were found between investigated groups. In contrast to serum concentrations, the expression of FCN2 gene (reported for the first time in ovarian sections) was significantly lower in women with OC in comparison with patients with NO but not with benign ovarian tumours. In case of FCN3 gene, its expression levels in OC group inversely correlated with serum ficolin-3 and were lower in comparison with controls.

Transient ST-segment elevation resembling acute myocardial infarction in a patient with a right secondary spontaneous pneumothorax.

A rare cause of ST-segment elevation mimicking myocardial infarction has been reported in the setting of acute pneumothorax. We present a middle-aged woman with a right-sided secondary pneumothorax who developed severe chest pain associated with ST-segment elevation suggestive of acute myocardial infarction. Symptoms resolved immediately after advancement of the dislodged chest drain. A subsequent coronary angiogram was normal. This case highlights an uncommon electrocardiographic alteration and discusses possible pathophysiological mechanisms.

Human L-ficolin (ficolin-2) and its clinical significance.

Human L-ficolin (P35, ficolin-2) is synthesised in the liver and secreted into the bloodstream where it is one of the major pattern recognition molecules of plasma/serum. Like other ficolins, it consists of a collagen-like tail region linked to a fibrinogen-related globular head; a basic triplet subunit arises via a collagen-like triple helix, and this then forms higher multimers (typically a 12-mer, Mr 400K). Unlike other ficolins, it has a complex set of binding sites arranged within an internal cleft enabling it to recognise a variety of molecular patterns including acetylated sugars and certain 1,3-ß-glucans. It is one of the few molecules known to activate the lectin pathway of complement. Recently, some disease association studies (at either the DNA or protein level) have implicated L-ficolin in innate immunity, where it might cooperate with pentraxins and collectins. Emerging lines of evidence point to a role for L-ficolin in respiratory immunity, where its affinity for Pseudomonas aeruginosa could be significant.

Factors of the lectin pathway of complement activation and their clinical associations in neonates.

This paper summarizes the data concerning soluble defense lectins (mannan-binding lectin, M-ficolin, L-ficolin, and H-ficolin) with the unique ability to activate complement and their associated serine proteases (MASPs) in neonates. The clinical importance of deficiencies of these immune factors is presented in aspects of perinatal mortality, premature births, and low birthweight. Prenatal serum concentrations of L-ficolin, H-ficolin, and MASP-2 (and probably M-ficolin) correlate with gestational age and birthweight. The relationship of serum MBL to gestational age is controversial. The MBL2 genotypes XA/O and O/O (associated with low-serum MBL) are associated with perinatal infections, whereas the high serum MBL-conferring A/A genotypes may be associated with prematurity. Low-serum L-ficolin concentrations, but not low-serum H-ficolin concentrations, are also associated with perinatal infections. Much of the literature is inconsistent, and the relationships reported so far require independent confirmation at both gene and protein levels. Our preliminary conclusion is that these soluble defense lectins play a protective role in the neonate, and that insufficiency of such factors contributes to the adverse consequences of prematurity and low birthweight.

Impact of mannose-binding lectin insufficiency on the course of cystic fibrosis: A review and meta-analysis.

Mannose-binding lectin (MBL) is an innate immune protein produced by the liver. MBL binds to glycoconjugates containing mannose, fucose or N-acetylglucosamine that are present in a wide variety of bacteria, viruses and fungi. Upon binding, MBL may active the lectin pathway of complement or directly opsonize organisms to enhance phagocytosis. MBL is primarily a serum protein but accumulates in the lung during acute inflammation. Recent evidence suggests an important role for MBL in a variety of infectious disorders. Cystic fibrosis (CF) is a multisystem disease caused by mutations in the gene encoding the CF transmembrane regulator (CFTR). The course of CF lung disease is highly variable even in patients with the same CFTR genotype, suggesting that other modulator genes are important for prognosis. MBL has been proposed as a possible modulator of clinical severity in CF. In this review and meta-analysis, we found that MBL2 genotypes associated with MBL insufficiency were associated with earlier acquisition of Pseudomonas aeruginosa (P < 0.0001), reduced pulmonary function among adult patients (P < 0.0001 for forced expiratory volume), and an increased rate of death or requirement for lung transplantation (odds ratio 3.69; P = 0.02). The available evidence therefore suggests that MBL insufficiency is associated with the severity of CF lung disease. The possible future prophylactic or therapeutic application of MBL replacement is discussed.

Polymorphisms of the FCN2 Gene 3'UTR Region and Their Clinical Associations in Preterm Newborns.

Ficolin-2 is regarded as an important innate immunity factor endowed with both lectin (carbohydrate recognition) qualities and ability to induce complement activation. The aim of this study was to investigate the association of the FCN2 3'-untranslated region (3'UTR) polymorphisms with ficolin-2 expression and perinatal complications in preterm neonates. The sequencing analysis allowed us to identify six 3'UTR polymorphisms with minor allele frequency (MAF) >1%: rs4521835, rs73664188, rs11103564, rs11103565, rs6537958 and rs6537959. Except for rs4521835, all adhered to Hardy-Weinberg expectations. Moreover, rs6537958 and rs6537959 were shown to be in perfect linkage disequilibrium (LD) with nine other genetic polymorphisms: rs7040372, rs7046516, rs747422, rs7847431, rs6537957, rs6537960, rs6537962, rs11462298 and rs7860507 together stretched on a distance of 1242 bp and very high LD with rs11103565. The 3'UTR region was shown to bind nuclear extract proteins. The polymorphisms at rs4521835 and rs73664188 were found to influence serum ficolin-2 concentration significantly. All polymorphisms identified create (together with exon 8 polymorphism, rs7851696) two haplotype blocks. Among 49 diplotypes (D1-D49) created from rs7851696 (G>T), rs4521835 (T>G), rs73664188 (T>C), rs11103564 (T>C), rs11103565 (G>A) and rs6537959 (T>A), twenty two occurred with frequency >1%. Two diplotypes: D13 (GTTTGT/GGTCGT) and D10 (GTTTGT/GGTCGA), were significantly more frequent among preterm neonates with early onset of infection and pneumonia, compared with newborns with no infectious complications (OR 2.69 and 2.81, respectively; both p<0.05). The minor (C) allele at rs73664188 was associated with an increased risk of very low (≤1500 g) birthweight (OR=1.95, p=0.042) but was associated with the opposite effect at rs11103564 (OR=0.11, p=0.005).

Two factors of the lectin pathway of complement, l-ficolin and mannan-binding lectin, and their associations with prematurity, low birthweight and infections in a large cohort of Polish neonates.

Ficolins and one collectin, mannan-binding lectin (MBL), are the only factors known to activate the lectin pathway (LP) of complement. There is considerable circumstantial evidence that MBL insufficiency can increase susceptibility to various infections and influence the course of several non-infectious diseases complicated by infections. Much less information is available concerning l-ficolin. We report the results of a prospective study to investigate any association between either MBL deficiency or l-ficolin deficiency with prematurity, low birthweight or perinatal infections in a large cohort of Polish neonates, representing an ethnically homogenous population (n=1832). Cord blood samples were analysed to determine mbl-2 gene variants, MBL concentrations and MBL-MASP-2 complex activities (MBL-dependent lectin pathway activity) as well as l-ficolin levels. Median concentrations of l-ficolin and MBL were 2500 and 1124 ng/ml, respectively, while median LP activity was 272 mU/ml. After genotyping, 60.6% of babies were mbl-2 A/A, 35.4% were A/O and 4% were O/O genotypes. We found relative l-ficolin deficiency to be associated with prematurity, low birthweight and infections. l-Ficolin concentration correlated with gestational age and with birthweight, independently of gestational age. Preterm deliveries (<38 weeks) occurred more frequently among neonates with low LP activity but not with those having low serum MBL levels. Similarly, no association of serum MBL deficiency with low birthweight was found, but there was a correlation between LP activity and birthweight. Genotypes conferring very low serum MBL concentrations were associated with perinatal infections, and high-MBL-conferring genotypes were associated with prematurity. Our findings suggest that l-ficolin participates in host defence during the perinatal period and constitute the first evidence that relative l-ficolin deficiency may contribute to the adverse consequences of prematurity. Some similar trends were found with facets of MBL deficiency, but the observed relationships were weaker and less consistent.

Branch retinal artery occlusion secondary to Bartonella henselae infection in a 13 year-old.

PURPOSE: To summarize the case of a 13 year-old boy diagnosed with a BRAO secondary to B. henselae infection. OBSERVATIONS: The patient presented with a sudden, unilateral, and painless scotoma. Fundoscopic findings and multimodal imaging were consistent with a BRAO with associated areas of intraretinal whitening along the involved artery. Upon further questioning, the patient reported having 15 cats at home. Antibodies were positive for B. henselae. The patient was treated with oral doxycycline 100 mg twice daily for 2 months with complete resolution of the retinal findings and the scotoma. CONCLUSIONS AND IMPORTANCE: B. henselae should be considered as a potential cause of retinitis and BRAO, even in pediatric-aged patients.

Hyperreflective Foci and Specks Are Associated with Delayed Rod-Mediated Dark Adaptation in Nonneovascular Age-Related Macular Degeneration.

PURPOSE: Hyperreflective foci (HRF) are OCT biomarkers for the progression of nonneovascular age-related macular degeneration (AMD) attributed to anteriorly migrated retinal pigment epithelial cells. We examined associations between rod- and cone-mediated vision and HRF plus smaller hyperreflective specks (HRS); we identified a histologic candidate for HRS. DESIGN: Cross-sectional study and histologic survey. PARTICIPANTS: Patients with healthy maculae (n = 34), early AMD (n = 26), and intermediate AMD (n = 41). METHODS: AMD severity was determined by color fundus photography. In OCT scans, HRF and HRS were counted manually. Vision tests probed cones (best-corrected visual acuity [VA], contrast sensitivity), mixed cones and rods (low-luminance VA, low-luminance deficit, mesopic light sensitivity), or rods (scotopic light sensitivity, rod-mediated dark adaptation [RMDA]). An online AMD histopathologic resource was reviewed. MAIN OUTCOME MEASURES: Vision in eyes assessed for HRF and HRS; histologic candidate for HRS. RESULTS: In 101 eyes of 101 patients, HRF and HRS were identified in 25 and 95 eyes, respectively, with good reliability. Hyperreflective foci were present but sparse in healthy eyes, infrequent in early AMD eyes, and frequent but highly variable among intermediate AMD eyes (mean±standard deviation [SD] number per eye, 0.1 ± 0.2, 0.2 ± 0.5, and 1.9 ± 3.4 for healthy, early AMD, and intermediate AMD eyes, respectively). Hyperreflective specks outnumbered HRF in all groups (mean±SD, 4.5 ± 3.2, 6.3 ± 5.8, and 19.4 ± 22.4, respectively). Delayed RMDA was associated strongly with more HRF and HRS (P < 0.0001). Hyperreflective foci also were associated with worse low-luminance VA (P = 0.0117). Hyperreflective specks were associated with worse contrast sensitivity (P = 0.0278), low-luminance VA (P = 0.0010), low-luminance deficit (P = 0.0031), and mesopic (P = 0.0018) and scotopic (P < 0.0001) sensitivity. By histologic analysis, cone lipofuscin was found in outer retinal layers of 25% of healthy aged eyes. CONCLUSIONS: Hyperreflective foci and HRS are markers of cellular activity associated with visual dysfunction, especially delayed RMDA, an AMD risk indicator assessing efficiency of retinoid resupply. Hyperreflective specks may represent lipofuscin translocating inwardly within cones. HRF and HRS may serve as structural end points in clinical trials targeting AMD stages earlier than atrophy expansion. These results should be confirmed in a larger sample.

Rapid group-, serotype-, and vaccine strain-specific identification of poliovirus isolates by real-time reverse transcription-PCR using degenerate primers and probes containing deoxyinosine residues.

We have adapted our previously described poliovirus diagnostic reverse transcription-PCR (RT-PCR) assays to a real-time RT-PCR (rRT-PCR) format. Our highly specific assays and rRT-PCR reagents are designed for use in the WHO Global Polio Laboratory Network for rapid and large-scale identification of poliovirus field isolates.

Mannan-binding lectin genotypes and genotype-phenotype relationships in a large cohort of Polish neonates.

Circulating mannan (or mannose)-binding lectin (MBL) is genetically determined. Low MBL concentrations are associated with certain point mutations in the human MBL2 gene. Here we report the full MBL2 genotypes of 1800 Polish neonates and relate individual genotypes to serum MBL and MBL-dependent activity of the lectin pathway of complement activation. The seven acknowledged common haplotypes were found, plus the uncommon LYPD haplotype, combining to form 33 genotypes in this population. As expected, a strong correlation existed between genotypes and serum MBL or lectin pathway activity, and the latter two entities correlated strongly with each other. However, serum MBL values varied up to greater than 90-fold within genotypes. Unexpectedly, higher lectin pathway activity was found in association with the P allele relative to the Q allele. These data from a large cohort of neonates, representing an ethnically homogenous population, suggest that the current knowledge of the genetics of MBL2 is inadequate to predict serum MBL concentration and MBL-dependent lectin pathway activity in individual subjects.

Bacteroides Spp. Blebitis, Keratitis, and Endophthalmitis Following Uncomplicated Trabeculectomy.

PURPOSE: To describe the first known case of Bacteroides spp. related blebitis, keratitis, and endophthalmitis following uncomplicated trabeculectomy. METHODS: This was a case report and literature review. CASE: A 63-year-old immunocompetent white male underwent uncomplicated trabeculectomy of his right eye. Two weeks later, a blebitis with adjacent keratitis was diagnosed, progressing over several days to endophthalmitis despite hourly topical fortified antibiotic therapy. Although gram stain and culture of the bleb surface, a conjunctival suture, the aqueous humor, and the vitreous were negative, topical real-time quantitative polymerase chain reaction testing disclosed the presence of Bacteroides spp. Following treatment with topical and intravitreal clindamycin and intravenous meropenem, all clinical evidence of infection resolved. Best spectacle-corrected visual acuity improved to 20/25 (0.8) subsequent to combined cataract extraction, intraocular lens implantation, and pars plana vitrectomy for persistent vitreous debris. CONCLUSIONS: Bacteroides may be a rare cause of postoperative blebitis, keratitis, and endophthalmitis. A favorable outcome may be attained, provided that an accurate diagnosis and effective treatment can be provided, which may be facilitated by real-time quantitative polymerase chain reaction in select cases.

Lectin-glycoconjugate interactions in health and disease.

It is increasingly being acknowledged that complex carbohydrates mediate a huge variety of cellular interactions, permitting and regulating recognition and signalling events. This is achieved by the enormous range and complexity of branched structures in glycoconjugates and the ability of carbohydrate-binding proteins (lectins) to decipher this 'glycocode'. Approx. 120 participants attended the 23rd International Lectin Meeting (Interlec-23) held at the Universities of Edinburgh (2 days) and Stirling (4 days) between 11 and 16 July 2008. These 'Interlecs' are truly international multi-disciplinary symposia, providing opportunities for scientists from different backgrounds, but with a common interest in some aspect of protein-carbohydrate interactions, to present their work in an informal and stimulating atmosphere. A major aim is always to induce cross-fertilization of ideas and concepts, and Interlec-23 was intended to have some bias towards lectins (galectins, collectins, selectins, siglecs etc.) and their ligands in human health and disease. Delegates from over 30 countries attended this meeting which was divided into seven oral sessions opened by a keynote speaker. This issue of Biochemical Society Transactions contains papers based on the keynote lectures and is therefore representative of the main themes of Interlec-23.

Is mannan-binding lectin (MBL) detectable on monocytes and monocyte-derived immature dendritic cells?

MBL (mannan-binding lectin; also called mannose-binding lectin) is a circulating C-type lectin with a collagen-like region synthesized mainly by the liver. MBL may influence susceptibility to infection in recipients of stem cell transplants, and it has even been suggested that the MBL status of a donor can influence the recipient's susceptibility to post-transplant infections. We have previously reported that MBL can be detected on human monocytes and monocyte-derived dendritic cells, based on detection using biotinylated anti-MBL, suggesting that those cells could synthesize MBL. If true, permanent MBL replacement therapy could be achieved by stem cell infusions. However, two other groups independently failed to find mbl-2-derived mRNA in monocytes. Therefore, to confirm or refute our previous observations, we used an alternative experimental strategy. Instead of using biotinylated antibody and labelled streptavidin, detection of surface MBL was attempted using MBL-specific primary antibodies (131-1, 131-10 and 131-11) followed by fluorescein-labelled anti-IgG, and controlled by the use of non-specific IgG as primary antibody. Monocytes were counterstained with anti-CD14-PE before FACS analysis. Adherent monocytes were also cultured for 48 h in serum-free medium or converted into immature dendritic cells by culture with IL-4 (interleukin-4) and GM-CSF (granulocyte/monocyte colony-stimulating factor). During FACS analysis, the dendritic cells were gated after counter-staining with anti-CD1a-PE. MBL was readily detected on the surface of fresh monocytes using all three specific anti-MBL monoclonal antibodies, but specific anti-MBL binding was greatly diminished after monocytes had been cultured for 2 days in serum-free medium. Moreover, we could not detect any MBL present on the surface of monocyte-derived dendritic cells. We therefore conclude that MBL is indeed present on the surface of fresh human monocytes. However, in view of the mRNA findings of others and our own previous observation that no secretion of MBL took place in culture, we presume that the surface-bound MBL is derived from autologous plasma and not synthesized by the cells. This conclusion is consistent with our in vivo findings in stem cell transplant patients which provided evidence against significant extra-hepatic production of serum MBL. It provides no ready explanation for the remarkable observation of Mullighan, Heatley, Doherty, Szabo, Grigg, Hughes, Schwarer, Szer, Tait, Bik To and Bardy [(2002) Blood 99, 3524-3529] that the presence of variant alleles of mbl-2 in stem cell donors can influence susceptibility to serious infections in their recipients.

Evaluation and comparison of the health status of Atlantic bottlenose dolphins from the Indian River Lagoon, Florida, and Charleston, South Carolina.

OBJECTIVE: To conduct health assessments and compare outcomes in 2 populations of Atlantic bottlenose dolphins. Design-Repeated cross-sectional study. ANIMALS: 171 Atlantic bottlenose dolphins. PROCEDURES: During June and August of 2003 through 2005, 89 dolphins from the Indian River Lagoon (IRL), Florida, and 82 dolphins from estuarine waters near Charleston, SC, were evaluated. A panel of 5 marine mammal veterinarians classified dolphins as clinically normal, possibly diseased, or definitely diseased on the basis of results of physical and ultrasonographic examinations, hematologic and serum biochemical analyses, and cytologic and microbiologic evaluations of gastric contents and swab specimens. RESULTS: Prevalence of dolphins classified as definitely diseased did not differ significantly between the IRL (32%) and Charleston (20%) sites. Proportions of dolphins classified as possibly diseased also did not differ. Lobomycosis was diagnosed in 9 dolphins from the IRL but in none of the dolphins from Charleston. Proportions of dolphins with orogenital papillomas did not differ significantly between the IRL (12%) and Charleston (7%) sites. From 2003 through 2005, the proportion classified as definitely diseased tripled among dolphins from the Charleston site but did not increase significantly among dolphins from the IRL. Dolphins from the Charleston site were more likely to have leukocytosis, lymphocytosis, and low serum concentrations of total protein and total J-globulins than were dolphins from the IRL. CONCLUSIONS AND CLINICAL RELEVANCE: High prevalences of diseased dolphins were identified at both sites; however, the host or environmental factors that contributed to the various abnormalities detected are unknown.

A solution method for the determination of cardiac potential distributions with an alternating current source.

A recently presented solution method for the bidomain model (Johnston et al. 2006), which involves the application of direct current for studying electrical potential in a slab of cardiac tissue, is extended here to allow the use of an applied alternating current. The advantage of using AC current, in a four-electrode method for determining cardiac conductivities, is that instead of using 'close' and 'wide' electrode spacings to make potential measurements, increasing the frequency of the AC current redirects a fraction of the current from the extracellular space into the intracellular space. The model is based on the work of Le Guyader et al. (2001), but is able to include the effects of the fibre rotation between the epicardium and the endocardium on the potentials. Also, rather than using a full numerical technique, the solution method uses Fourier series and a simple one dimensional finite difference scheme, which has the advantage of allowing the potentials to be calculated only at points, such as the measuring electrodes, where they are required. The new alternating current model, which includes intracellular capacitance, is used with a particular four-electrode configuration, to show that the potential measured is affected by changes in fibre rotation. This is significant because it indicates that it is necessary to include fibre rotation in models, which are to be used in conjunction with measuring arrays that are more complex than those involving simply surface probes or a single vertical probe.