Comprehensive biodiversity analysis via ultra-deep patterned flow cell technology: a case study of eDNA metabarcoding seawater.

The characterization of biodiversity is a crucial element of ecological investigations as well as environmental assessment and monitoring activities. Increasingly, amplicon-based environmental DNA metabarcoding (alternatively, marker gene metagenomics) is used for such studies given its ability to provide biodiversity data from various groups of organisms simply from analysis of bulk environmental samples such as water, soil or sediments. The Illumina MiSeq is currently the most popular tool for carrying out this work, but we set out to determine whether typical studies were reading enough DNA to detect rare organisms (i.e., those that may be of greatest interest such as endangered or invasive species) present in the environment. We collected sea water samples along two transects in Conception Bay, Newfoundland and analyzed them on the MiSeq with a sequencing depth of 100,000 reads per sample (exceeding the 60,000 per sample that is typical of similar studies). We then analyzed these same samples on Illumina's newest high-capacity platform, the NovaSeq, at a depth of 7 million reads per sample. Not surprisingly, the NovaSeq detected many more taxa than the MiSeq thanks to its much greater sequencing depth. However, contrary to our expectations this pattern was true even in depth-for-depth comparisons. In other words, the NovaSeq can detect more DNA sequence diversity within samples than the MiSeq, even at the exact same sequencing depth. Even when samples were reanalyzed on the MiSeq with a sequencing depth of 1 million reads each, the MiSeq's ability to detect new sequences plateaued while the NovaSeq continued to detect new sequence variants. These results have important biological implications. The NovaSeq found 40% more metazoan families in this environment than the MiSeq, including some of interest such as marine mammals and bony fish so the real-world implications of these findings are significant. These results are most likely associated to the advances incorporated in the NovaSeq, especially a patterned flow cell, which prevents similar sequences that are neighbours on the flow cell (common in metabarcoding studies) from being erroneously merged into single spots by the sequencing instrument. This study sets the stage for incorporating eDNA metabarcoding in comprehensive analysis of oceanic samples in a wide range of ecological and environmental investigations.

A rare case of Niemann-Pick disease type C without neurological involvement in a 66-year-old patient.

UNLABELLED: The case of a 66 year-old female - the oldest known living patient with Niemann-Pick disease type C (NP-C) who remains free of any neurological or psychiatric manifestations 18 years after presentation - is presented. An incidental finding of massive splenomegaly was detected during a routine pelvic ultrasound. The pathology report after splenectomy showed the presence of lipid-laden macrophages. Fibroblasts cultured in LDL-enriched medium revealed abnormal filipin staining consistent with cholesterol-filled vesicles and the rate of cholesterol esterification in response to stimulation of LDL-cholesterol uptake was significantly depressed at 6% of that seen in cells from normal controls, but at a level similar to that observed in an NP-C positive control. Molecular genetic testing later revealed a compound heterozygous mutant NP-C genotype comprising two previously described disease-causing mutations in the NPC1 gene, one in exon 8 (c.1133T>C [V378A]) and one in exon 13 (c.1990G>A [V664M]). These findings confirmed the diagnosis of NP-C. Only three patients with this disorder aged > 53 years have previously been reported, all of whom presented with neurological or neuropsychiatric manifestations. Our patient is the first reported NP-C patient, now in her seventh decade of life, who has to date only manifested splenomegaly. This case highlights the extreme clinical variability of NP-C, and the need to consider this disease in the differential diagnosis of organomegaly, even in the absence of neurological, psychiatric and related clinical signs. SYNOPSIS: An elderly female patient with confirmed NP-C and isolated splenomegaly has remained asymptomatic for neurological, cognitive, psychiatric or ophthalmologic abnormailities into her seventh decade of life.

'Sensitivity syndromes' related to radiation exposures.

The primary health effects of radiation are traditionally believed to result from cellular genetic damage. These effects are believed to result in a statistically detectable increase in the induction of cancer in exposed populations. A significant number of residents of areas affected by the Chernobyl disaster and workers involved in the clean-up ('liquidators') have reported debilitating physical illnesses that cannot be easily explained by a genetic effect. This paper presents results of a literature search that strongly suggests that a previously unrecognized neural pathway may be responsible for the induction of these debilities. In addition, a common link between radiation and chemical sensitivity syndromes may now be identified.

Use of the endometrial pipelle in the diagnosis of early abnormal gestations.

OBJECTIVE: To determine the sensitivity and specificity of the endometrial pipelle (Unimar, Wilton, CT) in detecting intrauterine chorionic villi, as compared to standard suction dilation and curettage (D and C). DESIGN: Prospective clinical study. SETTING: U. S. Navy Hospital near Tokyo, Japan. Small community hospital. PATIENT(S): All patients presenting for the surgical management of an early abnormal gestation were enrolled prospectively. The patient's age, gravidy, parity, estimated gestational age (EGA), diagnosis, preoperative beta human chorionic gonadotropin (betahCG) level, and preoperative transvaginal ultrasound (TVUS) data were recorded. INTERVENTION(S): An endometrial biopsy was performed in the operating room prior to standard suction dilatation and curettage (D and C). MAIN OUTCOME MEASURE(S): The presence of chorionic villi in the endometrial biopsy and curettage specimens were recorded for each patient. RESULT(S): The sensitivity of the endometrial pipelle in detecting intrauterine chorionic villi was 63% and the specificity was 80%. CONCLUSION(S): In the diagnosis of an early abnormal gestation, particularly in excluding an ectopic pregnancy, it would be ideal to replace the more invasive D and C with the outpatient endometrial biopsy in detecting intrauterine chorionic villi. However, the limited sensitivity of the endometrial biopsy limits its application in this clinical scenario. Our results compare with two previous studies and add to the limited patient database on this topic.

Some experiences in the estimation of parameters in non-linear differential equations.

The author describes a procedure developed by himself and his colleagues for obtaining estimates of the parameters of rate equations, together with information about confidence regions for the estimates. The program has been used successfully for processing results from the chemical engineering industry, with highly non-linear model systems, particularly since temperature was a variable, and the "rate constants" were non-linear combinations of other constants. In biochemical situations, in which investigations are almost always at constant temperature, the non-linearity should not be so extreme, and the procedure may well be capable of dealing with more than 5 to 7 parameters for which it is recommended.