The use of infrared spectroscopy and machine learning tools for detection of Meloidogyne infestations

Plant parasitic nematodes are generally soilborne pathogens that attack plants and cause economic losses in many crops. The infested plants show nonspecific symptoms or, often, are symptomless; therefore, diagnosis is performed by taking soil and root tissue samples. Here, we show that a combination of different infrared spectra analysis and machine learning algorithms can be used to detect plant parasitic nematode infestations before symptoms become visible, using leaves instead of roots and soil as samples. We found that tomato and guava plants infested with Meloidogyne enterorlobii produced different spectral patterns compared to uninfested plants. Using partial spectra from 1,450 to 900/cm as the "fingerprint region", principal component analyses indicated that after 5 (tomatoes) or 8 weeks (guava), plants with no visible symptoms of infestations were positively diagnosed. To improve the early detection response, we used machine learning modelling. A support vector machine (SVM) was used to obtain more robust, accurate models. The SVM model contained 34 support vectors, 17 for each level. The overall performance of the model was >97% and the total accuracy was significantly higher, demonstrating the absence of chance prediction. The best prediction of infestation was obtained at the second and fourth weeks for tomatoes and guavas, respectively, reducing the diagnostic time by half. The combined application of these techniques reduces the processing time from field to laboratory and shows enormous advantages by avoiding root and soil sampling.     

Fasciola hepatica: Histology of the testis in egg-producing adults of several laboratory-maintained isolates of flukes grown to maturity in cattle and sheep and in flukes from naturally infected hosts

A total of 8 calves approximately 6 months old and 22 lambs of similar age were infected with metacercariae of Fasciola hepatica of various laboratory-maintained isolates including: Cullompton (sensitive to triclabendazole) and Sligo, Oberon and Leon (reported as resistant to triclabendazole). Ten to 16 weeks after infection, flukes were harvested from these experimental animals and the histology of the testis tissue was examined in a representative sample of flukes from each population. Adult wild-type flukes were also collected from 5 chronically infected cattle and 7 chronically infected sheep identified at post-mortem inspection. The testis tissue of these flukes was compared with that of the various laboratory-maintained isolates. Whilst the testes of the wild-type, Oberon and Leon flukes displayed all the usual cell types associated with spermatogenesis in Fasciola hepatica (spermatogonia, spermatocytes, spermatids and mature sperm), the Cullompton flukes from both cattle and sheep showed arrested spermatogenesis, with no stages later than primary spermatocytes represented in the testis profiles. The presence of numerous eosinophilic apoptotic bodies and nuclear fragments suggested that meiotic division was anomalous and incomplete. In contrast to the wild-type flukes, no mature spermatozoa were present in the testes or amongst the shelled eggs in the uterus. A high proportion of the eggs collected from these flukes hatched to release normal-appearing miracidia after an appropriate incubation period, as indeed was the case with all isolates examined and the wild-type flukes. It is concluded that the eggs of Cullompton flukes are capable of development without fertilization, i.e. are parthenogenetic. The implications of this for rapid evolution of resistant clones following an anthelmintic selection event are discussed. Amongst the Sligo flukes examined, two subtypes were recognised, namely, those flukes with all stages of spermatogenesis and mature spermatozoa present in the testes (type 1), and those flukes with all stages of spermatogenesis up to spermatids present, but no maturing spermatozoa in the testes (type 2). Each sheep infected with the Sligo isolate had both type 1 (approximately 60%) and type 2 (approximately 40%) flukes present in the population. Spermatozoa were found amongst the eggs in the uterus in 64% of flukes and this did not necessarily reflect the occurrence of spermatozoa in the testis profiles of particular flukes, suggesting that cross-fertilization had occurred. The apparent disruption of meiosis in the spermatocytes of the Cullompton flukes is consistent with reports that Cullompton flukes are triploid (3n=30), whereas the Sligo and wild-type flukes are diploid (2n=20). In the Sligo flukes the populations are apparently genetically heterogenous, with a proportion of the flukes unable to produce fully formed spermatozoa perhaps because of a failure in spermiogenesis involving elongation of the nucleus during morphogenesis.     

Screening for resistance to Diaporthe toxica in lupins by estimation of phomopsins and glucoseamine in individual plants

Immunological and biochemical assays were developed for screening for resistance to Diaporthe toxica in individual plants of narrow-leafed lupins ( Lupinus angustifolius ). The former was an enzyme-linked immunosorbent assay (ELISA) for measuring phomopsin mycotoxins and the latter gave an estimation of glucoseamine in infected stem pieces. Stems of L. angustifolius seedlings were inoculated with conidia from D. toxica cultures and, as expected with this latent disease, remained symptomless for 21 days after inoculation. At this time, phomopsins were measured in excised stems that had been incubated for 6 or 8 days to allow mycelial growth from latent infection structures, thereby increasing the phomopsins to detectable levels in individual plants. The estimation of glucoseamine was carried out on the same stems that had been assayed for phomopsins. The method was based on the alkaline deacetylation of chitin to chitosan, the glucoseamine residues of which are de-aminated with nitrous acid, yielding an aldehyde which is determined colorimetrically. At six days after excision, both tests clearly distinguished the very resistant, resistant, intermediate and susceptible lines and they may be useful in large-scale resistance screening in lupin breeding programmes. The ELISA of phomopsins is easier to use and would be particularly useful in the elimination of susceptible plants and those plants expressing intermediate levels of resistance during early generations of the breeding programme.     

Growth and reproduction of the lion's paw scallop Nodipecten subnodosus in a suspended culture system at Guerrero Negro lagoon,Baja California Sur,Mexico

The lion's paw scallop, Nodipecten subnodosus (Sowerby) has considerable aquacultural potential due to its fast growth and large adductor muscle. Prior investigations throughout northwestern Mexico's littoral have reported highly variable growth rates; furthermore, no studies are available of the environment on growth and gametogenesis in this species under culture conditions. This investigation assesses the effect of food availability and temperature on the growth and gametogenesis of N. subnodosus in a suspended culture system at Guerrero Negro lagoon, Mexico. After 1 year of cultivation, N. subnodosus reached 69.13 mm in shell height (SH) (0.196 mm day^?1, 14 months old). Two significant growth spurts were observed: over the two first months of culture (August and September 2001, mean growth rate 0.4 mm day^?1) and in September 2002 (0.3 mm day^?1), both related to high temperatures and chlorophyll a concentrations. The onset of gametogenesis occurred in April 2002, with an increase in temperature (10‐month‐old scallops, 54.5 mm SH). The first spawning occurred in October and November (86.2 and 93 mm SH), with peak temperatures. These results, together with the analysis of previous reports, indicate that N. subnodosus has a higher preference for temperate areas; therefore, the Guerrero Negro lagoon appears to be a suitable site to culture this species.     

Karyotype analysis of interspecific hybrids between Haliotis rufescens and Haliotis discus HANNAI

Evidence of hybridization in Haliotis has been mainly supported by hatchery experiences and collection of wild hybrid abalones among several species from natural populations worldwide. However, despite the importance to understand the role of the hybridization process through Haliotidae evolution, and also its impact on the abalone aquaculture, genetic studies in hybrid abalones have been poorly developed. Herein, cytogenetic approach allows studying the genetic conformation in hybrid organisms at the chromosome level. This paper reports a quantitative karyotype analysis in Haliotis rufescens, Haliotis discus hannai and their interspecific hybrid. Thus, to characterize chromosome pairs and establish cytogenetic comparisons, chromosome banding with distamycin‐A/4,6‐diamidino‐2‐phenylindole fluorochromes and morphologic measurements were performed. The results showed that the hybrids are successfully viable and their karyotypes evidenced a conservative chromosome number of 2n=36. The karyo‐idiogram showed a high correspondence in chromosome pair morphology among the hybrids and their parental species, except for a single heteromorphic pair that corresponds to the chromosome 16 from H. rufescens andH. d. hannai respectively. The implications of the abalone hybrid viability derived from its chromosome composition are discussed.