A Case-control Study of Haemorrhagic Septicaemia in Buffaloes and Cattle in Karachi, Pakistan, in 2012.

A retrospective epidemiological case-control study was performed in Karachi, Pakistan, from January to April 2013. The owners of 217 dairy cattle and buffalo farms from six different locations in Karachi were interviewed. The aim of the study was to identify risk factors associated with the presence of haemorrhagic septicaemia (HS). Farms with a history of at least one instance of sudden death in a dairy animal during 2012 and a positive clinical HS diagnosis (made by local veterinarians) were defined as cases. Farms having no history of sudden deaths in 2012 were defined as controls. Univariable analyses were initially conducted, and factors with P ≤ 0.25 were offered to a multivariable logistic regression model to identify putative risk factors. The final multivariable logistic model contained five factors. Vaccination was found to be a protective factor (OR = 0.22) along with the length of time cattle were kept on farm (months). For every extra month cattle were kept, the odds of HS disease were reduced by a factor of 0.9. In contrast, for every extra animal in a herd, the risk of infection increased by a factor of 1.01. Supplying underground water and the presence of foot and mouth disease on the farm increased the risk by 2.90 and 2.37, respectively. To understand the epidemiology of HS in Karachi dairy herds, more in-depth research is required to study the risk and protective factors identified in this survey and to evaluate risk mitigation strategies, where possible.

A new Eimeria species parasitic in Isoodon obesulus (Marsupialia: Peramelidae) in Western Australia.

Feces from southern brown bandicoots, Isoodon obesulus, inhabiting the Perth metropolitan area were examined using fecal flotation and light microscopy, and were frequently found to contain oocysts (10/24; 42%). To enable formal description of the proposed new Eimeria species, i.e., Eimeria quenda n. sp., fecal oocysts from 1 juvenile male I. obesulus were allowed to sporulate in 2% potassium dichromate (K(2)Cr(2)O(7)) at room temperature. Sporulated oocysts are spheroidal to subspheroidal 24.5 × 23.6 (22.5-26.0 × 22.5-24.8) µm, with L/W ratio of 1.04 (1.00-1.13), lack a micropyle and oocyst residuum, and are contained within a smooth trilaminate oocyst wall 1.8 (1.6-2.0) µm thick. Sporocysts are ovoid, 12.6 × 9.2 (12.0-13.8 × 8.5-10.0) µm, with L/W ratio of 1.37 (1.20-1.53), have a sporocyst residuum, and 2 comma-shaped sporozoites, each containing 2 spheroidal refractile bodies. Sporulation takes 1-3 days at room temperature. This is the second formal description of an Eimeria species parasitic in the order Peramelemorphia.

Nuclear DNA amounts in angiosperms: targets, trends and tomorrow.

BACKGROUND AND AIMS: The amount of DNA in an unreplicated gametic chromosome complement is known as the C-value and is a key biodiversity character of fundamental significance with many practical and predictive uses. Since 1976, Bennett and colleagues have assembled eight compilations of angiosperm C-values for reference purposes and subsequently these have been pooled into the Angiosperm DNA C-values Database (http://data.kew.org/cvalues/). Since the last compilation was published in 2005, a large amount of data on angiosperm genome size has been published. It is therefore timely to bring these data together into a ninth compilation of DNA amounts. Scope The present work lists DNA C-values for 2221 species from 151 original sources (including first values for 1860 species not listed in previous compilations). Combining these data with those published previously shows that C-values are now available for 6287 angiosperm species. KEY FINDINGS: Analysis of the dataset, which is by far the largest of the nine compilations published since 1976, shows that angiosperm C-values are now being generated at the highest rate since the first genome sizes were estimated in the 1950s. The compilation includes new record holders for the smallest (1C = 0·0648 pg in Genlisea margaretae) and largest (1C = 152·23 pg in Paris japonica) genome sizes so far reported, extending the range encountered in angiosperms to nearly 2400-fold. A review of progress in meeting targets set at the Plant Genome Size meetings shows that although representation for genera, geographical regions and some plant life forms (e.g. island floras and parasitic plants) has improved, progress to increase familial representation is still slow. In terms of technique it is now clear that flow cytometry is soon likely to become the only method available for plant genome size estimations. Fortunately, this has been accompanied by numerous careful studies to improve the quality of data generated using this technique (e.g. design of new buffers, increased awareness and understanding of problems caused by cytosolic inhibitors). It is also clear that although the speed of DNA sequencing continues to rise dramatically with the advent of next-generation and third-generation sequencing technologies, 'complete genome sequencing' projects are still unable to generate accurate plant genome size estimates.

Cutaneous papillomatosis and carcinomatosis in the Western barred bandicoot (Perameles bougainville).

A progressive wart-like syndrome in both captive and wild populations of the Western barred bandicoot (WBB) is hindering conservation efforts to prevent the extinction of this endangered marsupial. In this study, 42 WBBs exhibiting the papillomatosis and carcinomatosis syndrome were examined. The disease was characterized by multicentric proliferative lesions involving cutaneous and mucosal surfaces, which were seen clinically to increase in size with time. Grossly and histologically the smaller skin lesions resembled papillomas, whereas the larger lesions were most commonly observed to be squamous cell carcinomas. Large amphophilic intranuclear inclusion bodies were observed in hyperplastic conjunctival lesions of 8 WBBs under light microscopy. Conjunctival lesions from 2 WBBs examined using transmission electron microscopy contained a crystalline array of spherical electron-dense particles of 45-nm diameter, within the nucleus of conjunctival epithelial cells, consistent with a papillomavirus or polyomavirus. Conjunctival samples from 3 bandicoots that contained intranuclear inclusion bodies also demonstrated a positive immunohistochemical reaction after indirect immunohistochemistry for papillomavirus structural antigens. Ultrastructural and/or immunohistochemical evidence of an etiologic agent was not identified in the nonconjunctival lesions examined. Here we describe the gross, histopathologic, ultrastructural, and immunohistochemical findings of a papillomatosis and carcinomatosis syndrome recently identified in the WBB.

Klossiella quimrensis (Apicomplexa: Klossiellidae) causes renal coccidiosis in western barred bandicoots Perameles bougainville (Marsupialia: Peramelidae) in Western Australia.

Previous studies have described a range of Klossiella species parasitic in marsupial hosts. Klossiella quimrensis is the etiologic agent of renal coccidiosis in the peramelid marsupial hosts Isoodon obesulus and Perameles gunnii in Eastern Australia, but there is no previous report of klossiellosis in Western Australian peramelids. This study describes klossiellosis diagnosed by histology of renal tissue sections collected during necropsy of 20 Perameles bougainville between 2000 and 2005. Sporonts, sporoblasts, and macrogametes were identified within parasitophorous vacuoles of epithelial cells located near the renal corticomedullary junction. The prevalence of renal coccidiosis in P. bougainville diagnosed by renal histology is estimated at 30%. Only a single unsporulated sporocyst was detected by examination of cystocentesis-collected urine, indicating that microscopic evaluation of urine samples is an insensitive diagnostic test for detection of K. quimrensis in P. bougainville. This infection in P. bougainville is indirectly associated with mild multifocal interstitial lymphohistiocytic nephritis and is likely to be only minimally pathogenic in otherwise healthy individuals. Our study also extends the host and geographic range of K. quimrensis to include P. bougainville and Western Australia.

Homicide of children aged 0-4 years, 2003-04: results from the National Violent Death Reporting System.

INTRODUCTION: To better understand, and ultimately prevent, infant/child homicide, it is imperative to more thoroughly elucidate the circumstances and conditions related to such instances. Data were obtained from the US National Violent Death Reporting System (NVDRS) to illuminate circumstances related to homicide among children aged 4 years or less, and to identify demographic groups which may be at increased risk. METHODS: The NVDRS is an active surveillance system that provides comprehensive information on all violent deaths that occur within participating states within the US. Standard statistical tests were conducted to determine homicide rates among children ages 0-4 across states that provided data for both 2003 and 2004 (Alaska, Maryland, Massachusetts, New Jersey, Oregon, South Carolina, and Virginia). These data were further used to investigate infant/child homicides by race, gender, and other relevant circumstances (for example, victim-suspect relationship, weapon type, and location of homicide). A Poisson regression model was fitted to the sample data to investigate the multivariate relationship between the infant/child homicide rate and available demographic information. RESULTS: The 2003 homicide rate for children ages 0-4 was 3.0 per 100,000 population. The 2004 homicide rate was 2.5 per 100,000 population. African Americans were 4.2 times as likely as whites to be victims of homicide. Suspects were commonly parents/caregivers. The vast majority of infant/child homicides occurred in houses or apartments, using weapons that include household objects. CONCLUSION: Homicides of infants and young children are most often committed in the home, by parents/caregivers, using "weapons of opportunity". This suggests that the risk of infant/child homicide is greatest within the primary care giving environment. Moreover, the use of "weapons of opportunity" may be indicative of maladaptive stress responses. Prevention and intervention strategies to reduce infant/child homicide should target the home environment and attend to maladaptive stress responses.

A new eimeria species parasitic in western barred bandicoots, Perameles bougainville (Marsupialia: Peramelidae), in western Australia.

Feces from western barred bandicoots, Perameles bougainville, examined during routine monitoring of captive breeding colonies and wild populations were frequently found to contain oocysts. Fecal oocysts from 1 individual housed at Kanyana Wildlife Rehabilitation Centre were allowed to sporulate in 2% potassium dichromate (K2Cr2O7) at room temperature. Sporulated oocysts are subspheroidal 18.8 X 17.9 (16.9-21.0 x 16.0-19.9) microm, with length/width (L/W) ratio of 1.05 (1.00-1.15), lack a micropyle and oocyst residuum, but they usually have a polar granule within a smooth trilaminate oocyst wall 1.0 (0.7-1.3) microm thick. Sporocysts are ovoid, 9.1 x 7.0 (8.1-10.8 x 6.1-8.6) microm, with L/W ratio of 1.32 (1.04-1.51), have a Stieda body, sporocyst residuum, and 2 comma-shaped sporozoites, each containing 2 spheroidal refractile bodies. Sporulation takes 2-5 days at room temperature. This is the first formal description of an Eimeria species parasitic in the order Peramelemorphia.

First nuclear DNA amounts in more than 300 angiosperms.

BACKGROUND AND AIMS: Genome size (DNA C-value) data are key biodiversity characters of fundamental significance used in a wide variety of biological fields. Since 1976, Bennett and colleagues have made scattered published and unpublished genome size data more widely accessible by assembling them into user-friendly compilations. Initially these were published as hard copy lists, but since 1997 they have also been made available electronically (see the Plant DNA C-values database http://www.kew.org/cval/homepage.html). Nevertheless, at the Second Plant Genome Size Meeting in 2003, Bennett noted that as many as 1000 DNA C-value estimates were still unpublished and hence unavailable. Scientists were strongly encouraged to communicate such unpublished data. The present work combines the databasing experience of the Kew-based authors with the unpublished C-values produced by Zonneveld to make a large body of valuable genome size data available to the scientific community. METHODS: C-values for angiosperm species, selected primarily for their horticultural interest, were estimated by flow cytometry using the fluorochrome propidium iodide. The data were compiled into a table whose form is similar to previously published lists of DNA amounts by Bennett and colleagues. KEY RESULTS AND CONCLUSIONS: The present work contains C-values for 411 taxa including first values for 308 species not listed previously by Bennett and colleagues. Based on a recent estimate of the global published output of angiosperm DNA C-value data (i.e. 200 first C-value estimates per annum) the present work equals 1.5 years of average global published output; and constitutes over 12 % of the latest 5-year global target set by the Second Plant Genome Size Workshop (see http://www.kew.org/cval/workshopreport.html). Hopefully, the present example will encourage others to unveil further valuable data which otherwise may lie forever unpublished and unavailable for comparative analyses.

Plant genome size research: a field in focus.

This Special Issue contains 18 papers arising from presentations at the Second Plant Genome Size Workshop and Discussion Meeting (hosted by the Royal Botanic Gardens, Kew, 8-12 September, 2003). This preface provides an overview of these papers, setting their key contents in the broad framework of this highly active field. It also highlights a few overarching issues with wide biological impact or interest, including (1) the need to unify terminology relating to C-value and genome size, (2) the ongoing quest for accurate gold standards for accurate plant genome size estimation, (3) how knowledge of species' DNA amounts has increased in recent years, (4) the existence, causes and significance of intraspecific variation, (5) recent progress in understanding the mechanisms and evolutionary patterns of genome size change, and (6) the impact of genome size knowledge on related biological activities such as genetic fingerprinting and quantitative genetics. The paper offers a vision of how increased knowledge and understanding of genome size will contribute to holisitic genomic studies in both plants and animals in the next decade.

Nuclear DNA amounts in angiosperms: progress, problems and prospects.

BACKGROUND: The nuclear DNA amount in an unreplicated haploid chromosome complement (1C-value) is a key diversity character with many uses. Angiosperm C-values have been listed for reference purposes since 1976, and pooled in an electronic database since 1997 (http://www.kew.org/cval/homepage). Such lists are cited frequently and provide data for many comparative studies. The last compilation was published in 2000, so a further supplementary list is timely to monitor progress against targets set at the first plant genome size workshop in 1997 and to facilitate new goal setting. SCOPE: The present work lists DNA C-values for 804 species including first values for 628 species from 88 original sources, not included in any previous compilation, plus additional values for 176 species included in a previous compilation. CONCLUSIONS: 1998-2002 saw striking progress in our knowledge of angiosperm C-values. At least 1700 first values for species were measured (the most in any five-year period) and familial representation rose from 30 % to 50 %. The loss of many densitometers used to measure DNA C-values proved less serious than feared, owing to the development of relatively inexpensive flow cytometers and computer-based image analysis systems. New uses of the term genome (e.g. in 'complete' genome sequencing) can cause confusion. The Arabidopsis Genome Initiative C-value for Arabidopsis thaliana (125 Mb) was a gross underestimate, and an exact C-value based on genome sequencing alone is unlikely to be obtained soon for any angiosperm. Lack of this expected benchmark poses a quandary as to what to use as the basal calibration standard for angiosperms. The next decade offers exciting prospects for angiosperm genome size research. The database (http://www.kew.org/cval/homepage) should become sufficiently representative of the global flora to answer most questions without needing new estimations. DNA amount variation will remain a key interest as an integrated strand of holistic genomics.

Evolution of DNA amounts across land plants (embryophyta).

BACKGROUND AND AIMS: DNA C-values in land plants (comprising bryophytes, lycophytes, monilophytes, gymnosperms and angiosperms) vary approximately 1000-fold from approx. 0.11 to 127.4 pg. To understand the evolutionary significance of this huge variation it is essential to evaluate the phylogenetic component. Recent increases in C-value data (e.g. Plant DNA C-values database; release 2.0, January 2003; http://www.rbgkew.org.uk/cval/homepage.html) together with improved consensus of relationships between and within land plant groups makes such an analysis timely. METHODS: Insights into the distribution of C-values in each group of land plants were gained by superimposing available C-value data (4119 angiosperms, 181 gymnosperms, 63 monilophytes, 4 lycophytes and 171 bryophytes) onto phylogenetic trees. To enable ancestral C-values to be reconstructed for clades within land plants, character-state mapping with parsimony and MacClade was also applied. KEY RESULTS AND CONCLUSIONS: Different land plant groups are characterized by different C-value profiles, distribution of C-values and ancestral C-values. For example, the large ( approximately 1000-fold) range yet strongly skewed distribution of C-values in angiosperms contrasts with the very narrow 12-fold range in bryophytes. Further, character-state mapping showed that the ancestral genome sizes of both angiosperms and bryophytes were reconstructed as very small (i.e. < or =1.4 pg) whereas gymnosperms and most branches of monilophytes were reconstructed with intermediate C-values (i.e. >3.5, <14.0 pg). More in-depth analyses provided evidence for several independent increases and decreases in C-values; for example, decreases in Gnetaceae (Gymnosperms) and heterosperous water ferns (monilophytes); increases in Santalales and some monocots (both angiosperms), Pinaceae, Sciadopityaceae and Cephalotaxaceae (Gymnosperms) and possibly in the Psilotaceae + Ophioglossaceae clade (monilophytes). Thus, in agreement with several focused studies within angiosperm families and genera showing that C-values may both increase and decrease, it is apparent that this dynamic pattern of genome size evolution is repeated on a broad scale across land plants.

Pseudophysaloptera enigmatica sp. nov. (Nematoda: Physalopteridae) from Dactylopsila trivirgata Gray, 1858 (Marsupialia: Petauridae) in Australia, with a review of the genus.

The nematode Pseudophysaloptera enigmatica sp. nov. is described from the stomach of Dactylopsila trivirgata Gray, 1858, in Queensland, Australia. The new species is distinguished from congeners by its size and features present on the tail of the male. P. enigmatica sp. nov. is considerably larger than virtually all congeners, and can be differentiated by the presence of spicules and the shape and distribution of caudal papillae in the male tail. This is the first record of the genus in Australia and in marsupials. A review of the genus is provided.

Telomere variability in the monocotyledonous plant order Asparagales.

A group of monocotyledonous plants within the order Asparagales, forming a distinct clade in phylogenetic analyses, was reported previously to lack the 'typical' Arabidopsis-type telomere (TTTAGGG)(n). This stimulated us to determine what has replaced these sequences. Using slot-blot and fluorescent in situ hybridization (FISH) to species within this clade, our results indicate the following. 1. The typical Arabidopsis-type telomeric sequence has been partly or fully replaced by the human-type telomeric sequence (TTAGGG)(n). Species in Allium lack the human-type variant. 2. In most cases the human variant occurs along with a lower abundance of two or more variants of the minisatellite sequences (of seven types evaluated), usually these being the consensus telomeric sequence of Arabidopsis, Bombyx (TTAGG)(n) and Tetrahymena (TTGGGG)(n). FISH shows that the variants can occur mixed together at the telomere. 3. Telomerases generate products with a 6 base pair periodicity and when sequenced they reveal predominantly a reiterated human-type motif. These motifs probably form the 'true telomere' but the error rate of motif synthesis is higher compared with 'typical' plant telomerases. The data indicate that the Asparagales clade is unified by a mutation resulting in a switch from synthesis of Arabidopsis-like telomeres to a low-fidelity synthesis of human-like telomeres.

Loss and recovery of Arabidopsis-type telomere repeat sequences 5'-(TTTAGGG)(n)-3' in the evolution of a major radiation of flowering plants.

Fluorescent in situ hybridization and Southern blotting were used for showing the predominant absence of the Arabidopsis-type telomere repeat sequence (TRS) 5'-(TTTAGGG)(n)-3' (the 'typical' telomere) in a monocot clade which comprises up to 6300 species within Asparagales. Initially, two apparently disparate genera that lacked the typical telomere were identified. Here, we used the new angiosperm phylogenetic classification for predicting in which other related families such telomeres might have been lost. Our data revealed that 16 species in 12 families of Asparagales lacked typical telomeres. Phylogenetically, these were clustered in a derived clade, thereby enabling us to predict that the typical telomere was lost, probably as a single evolutionary event, following the divergence of Doryanthaceae ca. 80--90 million years ago. This result illustrates the predictive value of the new phylogeny, as the pattern of species lacking the typical telomere would be considered randomly placed against many previous angiosperm taxonomies. Possible mechanisms by which chromosome end maintenance could have evolved in this group of plants are discussed. Surprisingly, one genus, Ornithogalum (Hyacinthaceae), which is central to the group of plants that have lost the typical telomere, appears to have regained the sequences. The mechanism(s) by which such recovery may have occurred is unknown, but possibilities include horizontal gene transfer and sequence reamplification.

Modified technique of haptic externalization for scleral fixation of dislocated posterior chamber lens implants.

PURPOSE: To describe a modified technique of haptic externalization during repositioning of dislocated posterior chamber lens implants, which facilitates placement of scleral fixation sutures around the haptic for implant stabilization. METHODS: We describe a technique of repositioning a dislocated posterior chamber implant with scleral fixation sutures, which uses a small, clear corneal incision for externalization of the haptic. After a loop of 10-0 Prolene suture (Ethicon, Inc., Somerville, New Jersey) is placed around the externalized haptic, the sutures are retrieved through a sclerotomy 1.0 mm posterior to the limbus. The haptic is reimplanted into the ciliary sulcus. A separate scleral fixation bite closes the sclerotomy, and it is tied to the 10-0 Prolene sutures looped around the haptic. RESULTS: The dislocated implant was stable and fixated in good position 5 months after surgery using this technique. CONCLUSION: This externalization technique minimizes the extensive intraocular manipulations necessary to create a suture loop around a haptic of a dislocated implant. The clear corneal incision allows for clear visualization during externalization of the haptic.

Ribosomal DNA evolution and phylogeny in Aloe (Asphodelaceae).

All Aloe taxa (∼400 species) share a conserved bimodal karyotype with a basic genome of four large and three small submetacentric/acrocentric chromosomes. We investigated the physical organization of 18S-5.8S-26S and 5S ribosomal DNA (rDNA) using fluorescent in situ hybridization (FISH) to 13 Aloe species. The organization was compared with a phylogenetic tree of 28 species (including the 13 used for FISH) constructed by sequence analysis of the internal transcribed spacer (ITS) of 18S-5.8S-26S rDNA. The phylogeny showed little divergence within Aloe, although distinct, well-supported clades were found. FISH analysis of 5S rDNA distribution showed a similar interstitial location on a large chromosome in all species examined. In contrast, the distribution of 18S-5.8S-26S rDNA was variable, with differences in number, location, and size of loci found between species. Nevertheless, within well-supported clades, all species had the same organizational patterns. Thus, despite the striking stability of karyotype structure and location of 5S rDNA, the distribution of 18S-5.8S-26S rDNA is not so constrained and has clearly changed during Aloe speciation.

Aloe L.--a second plant family without (TTTAGGG)n telomeres.

The physical ends of chromosomes are protected and stabilised by telomeres. The sequence of telomeric DNA normally consists of a simple repeating unit that is conserved in many organisms. Most plants examined have been shown to possess Arabidopsis-type telomeres consisting of many repeat copies of the sequence 5'-TTTAGGG-3'. Using fluorescent in situ hybridisation, slot blotting and the asymmetric polymerase chain reaction we demonstrate an absence of Arabidopsis-type telomeres in the genus Aloe (family Asphodelaceae). The only other plant genera so far reported without such telomeres are Allium, Nothoscordum, and Tulbaghia (family Alliaceae). As these genera and Aloe are petaloid monocots in the Asparagales, it is suggested that an absence of Arabidopsis-type telomeres may be characteristic of this related group of plants.

Genome structure and evolution in the allohexaploid weed Avena fatua L. (Poaceae).

Allohexaploid wild oat, Avena fatua L. (Poaceae; 2n = 6x = 42), is one of the world's worst weeds, yet unlike some of the other Avena hexaploids, its genomic structure has been relatively little researched. Consequently, in situ hybridisation was carried out on one accession of A. fatua using an 18S-25S ribosomal DNA (rDNA) sequence and genomic DNA from A. strigosa (AA-genome diploid) and A. clauda (CC-genome diploid) as probes. Comparing these results with those for other hexaploids studied previously: (i) confirmed that the genomic composition of A. fatua was similar to the other hexaploid Avena taxa (i.e., AACCDD), (ii) identified major sites of rDNA on three pairs of A/D-genome chromosomes, in common with other Avena hexaploids, and (iii) revealed eight chromosome pairs carrying intergenomic translocations between the A/D- and C-genomes in the accession studied. Based on karyotype structure, the identity of some of these recombinant chromosomes was proposed, and this showed that some of these could be divided into two types, (i) those common to all hexaploid Avena species analysed (3 translocations) and (ii) one translocation in this A. fatua accession not previously observed in reports on other hexaploid Avena species. If this translocation is found to be unique to A. fatua, then this information, combined with more traditional morphological data, will add support to the view that A. fatua is genetically distinct from other hexaploid Avena species and thus should retain its full specific status.

Reference standards for determination of DNA content of plant nuclei.

Flow cytometry was used to compare 14 potential reference standards for plant DNA content determination. Both chicken and plant internal standards were used, as were propidium iodide (PI) and 4'-6-diamidino-2-phenylindole (DAPI) as fluorochromes. Means and standard errors of the means are presented for the 14 potential reference standards, and the means are compared to those obtained by Feulgen densitometry. Five species are recommended as an initial set of international standards for future plant DNA content determinations: Sorghum bicolor cv. Pioneer 8695 (2C = 1.74 pg), Pisum sativum cv. Minerva Maple (2C = 9.56 pg), Hordeum vulgare cv. Sultan (2C = 11.12 pg), Vicia faba (2C = 26.66 pg), and Allium cepa cv. Ailsa Craig (2C = 33.55 pg). It is recommended that the reference standard of choice be one with 2C and 4C nuclear DNA content peaks similar to, but not overlapping, the 2C and 4C peaks of the target species. We recommend PI as the fluorochrome of choice for flow cytometric determination of plant DNA content. DAPI should be used only if the estimated DNA value is corroborated by using a second stain that has no bias for AT- or GC-rich sequences within genomes.