Survival, growth and tag retention of juvenile European eel (Anguilla anguilla L.) with implanted 12 mm passive integrated transponder tags and acoustic tags.

To evaluate the efficiency of tagging juvenile European eels with implanted 12 mm passive integrated transponder (PIT) tags or Eel/Lamprey acoustic transmitters (ELATs), the authors studied tag retention, survival and growth of eels (7-25 g). Experimental eels were obtained from an eel farm, tagged and then released in a series of shallow dug-out ponds with a surface area of c. 200 m2 . Tagged and control eels were distributed evenly, with 50 tagged and 50 control eels in each of four ponds, giving a total of 200 tagged and 200 control eels mixed. After 76 days, the ponds were drained, and eels were sampled and measured. A total of 344 eels (86%) were recaptured, indicating high survival. Tag retention was 99% as only one of the recaptured PIT-tagged eels had lost the tag and none of the ELAT tagged. The results demonstrated that tagging juvenile eels >16 cm with these small tags is indeed feasible. The growth of tagged and control fish was differentiated but generally low in length and negative in mass but did not differ between the three groups.

Observations on the use of passive integrated transponder (PIT) tags in teleosts and elasmobranchs at a public aquarium, 728 cases, 2007-2020.

Between 2007 and 2020 at New England Aquarium, Boston, MA, USA, we implanted passive integrated transponder (PIT) tags into 728 fish representing 105 teleost and elasmobranch species to identify animals as individuals. At the time of retrospective data analysis, mean longevity interval (median, range) after tag placement for animals that remained alive (n = 236) was 4.7 years (4.5, 0.3-13.8). Mean interval (median, range) between tag placement and death (n = 317) was 2.1 years (1.6, 0-11.2); and mean interval (median, range) between tag placement and transfer to other facilities (n = 175) was 2.5 years (3.1, 0.1-9.3). Possible adverse effects of tagging were extremely rare. Using the described methods, the equipment cost for every 10 PIT tag implantations was $2.83. PIT tag implantation in fishes is a safe and cost effective method to identify individuals, providing an opportunity to accumulate valuable data regarding individual longevity, welfare, basic demographics, and outcome of medical management. PIT tag implantation is recommended as a routine aspect of acquisition, quarantine, and medical management of fish under human care.

Development and validation of a 1 K sika deer (Cervus nippon) SNP Chip.

BACKGROUND: China is the birthplace of the deer family and the country with the most abundant deer resources. However, at present, China's deer industry faces the problem that pure sika deer and hybrid deer cannot be easily distinguished. Therefore, the development of a SNP identification chip is urgently required. RESULTS: In this study, 250 sika deer, 206 red deer, 23 first-generation hybrid deer (F1), 20 s-generation hybrid deer (F2), and 20 third-generation hybrid deer (F3) were resequenced. Using the chromosome-level sika deer genome as the reference sequence, mutation detection was performed on all individuals, and a total of 130,306,923 SNP loci were generated. After quality control filtering was performed, the remaining 31,140,900 loci were confirmed. From molecular-level and morphological analyses, the sika deer reference population and the red deer reference population were established. The Fst values of all SNPs in the two reference populations were calculated. According to customized algorithms and strict screening principles, 1000 red deer-specific SNP sites were finally selected for chip design, and 63 hybrid individuals were determined to contain red deer-specific SNP loci. The results showed that the gene content of red deer gradually decreased in subsequent hybrid generations, and this decrease roughly conformed to the law of statistical genetics. Reaction probes were designed according to the screening sites. All candidate sites met the requirements of the Illumina chip scoring system. The average score was 0.99, and the MAF was in the range of 0.3277 to 0.3621. Furthermore, 266 deer (125 sika deer, 39 red deer, 56 F1, 29 F2,17 F3) were randomly selected for 1 K SNP chip verification. The results showed that among the 1000 SNP sites, 995 probes were synthesized, 4 of which could not be typed, while 973 loci were polymorphic. PCA, random forest and ADMIXTURE results showed that the 1 K sika deer SNP chip was able to clearly distinguish sika deer, red deer, and hybrid deer and that this 1 K SNP chip technology may provide technical support for the protection and utilization of pure sika deer species resources. CONCLUSION: We successfully developed a low-density identification chip that can quickly and accurately distinguish sika deer from their hybrid offspring, thereby providing technical support for the protection and utilization of pure sika deer germplasm resources.

Assessing DNA for fish identifications from reference collections: the good, bad and ugly shed light on formalin fixation and sequencing approaches.

Natural history collections are repositories of biodiversity and are potentially used by molecular ecologists for comparative taxonomic, phylogenetic, biogeographic and forensic purposes. Specimens in fish collections are preserved using a combination of methods with many fixed in formalin and then preserved in ethanol for long-term storage. Formalin fixation damages DNA, thereby limiting genetic analyses. In this study, the authors compared the DNA barcoding and identification success for frozen and formalin-fixed tissues obtained from specimens in the CSIRO Australian National Fish Collection. They studied 230 samples from fishes (consisting of >160 fish species). An optimized formalin-fixed, paraffin-embedded DNA extraction method resulted in usable DNA from degraded tissues. Four mini barcoding assays of the mitochondrial DNA (mtDNA) were characterized with Sanger and Illumina amplicon sequencing. In the good quality DNA (without exposure to formalin), up to 88% of the specimens were correctly matched at the species level using the cytochrome oxidase subunit 1 (COI) mini barcodes, whereas up to 58% of the specimens exposed to formalin for less than 8 weeks were correctly identified to species. In contrast, 16S primers provided higher amplification success with formalin-exposed tissues, although the COI gene was more successful for identification. Importantly, the authors found that DNA of a certain size and quality can be amplified and sequenced despite exposure to formalin, and Illumina sequencing provided them with greater power of resolution for taxa identification even when there was little DNA present. Overall, within parameter constraints, this study highlights the possibilities of recovering DNA barcodes for identification from formalin-fixed fish specimens, and the authors provide guidelines for when successful identification could be expected.

Longer is Not Always Better: Optimizing Barcode Length for Large-Scale Species Discovery and Identification.

New techniques for the species-level sorting of millions of specimens are needed in order to accelerate species discovery, determine how many species live on earth, and develop efficient biomonitoring techniques. These sorting methods should be reliable, scalable, and cost-effective, as well as being largely insensitive to low-quality genomic DNA, given that this is usually all that can be obtained from museum specimens. Mini-barcodes seem to satisfy these criteria, but it is unclear how well they perform for species-level sorting when compared with full-length barcodes. This is here tested based on 20 empirical data sets covering ca. 30,000 specimens (5500 species) and six clade-specific data sets from GenBank covering ca. 98,000 specimens ($>$20,000 species). All specimens in these data sets had full-length barcodes and had been sorted to species-level based on morphology. Mini-barcodes of different lengths and positions were obtained in silico from full-length barcodes using a sliding window approach (three windows: 100 bp, 200 bp, and 300 bp) and by excising nine mini-barcodes with established primers (length: 94-407 bp). We then tested whether barcode length and/or position reduces species-level congruence between morphospecies and molecular operational taxonomic units (mOTUs) that were obtained using three different species delimitation techniques (Poisson Tree Process, Automatic Barcode Gap Discovery, and Objective Clustering). Surprisingly, we find no significant differences in performance for both species- or specimen-level identification between full-length and mini-barcodes as long as they are of moderate length ($>$200 bp). Only very short mini-barcodes (<200 bp) perform poorly, especially when they are located near the 5$^\prime$ end of the Folmer region. The mean congruence between morphospecies and mOTUs was ca. 75% for barcodes $>$200 bp and the congruent mOTUs contain ca. 75% of all specimens. Most conflict is caused by ca. 10% of the specimens that can be identified and should be targeted for re-examination in order to efficiently resolve conflict. Our study suggests that large-scale species discovery, identification, and metabarcoding can utilize mini-barcodes without any demonstrable loss of information compared to full-length barcodes. [DNA barcoding; metabarcoding; mini-barcodes; species discovery.].

A field evaluation of long-term effects of PIT tagging.

Passive integrated transponder (PIT)-tagging is commonly used in behavioural studies of fish, although long-term evaluations of effects from tagging under natural conditions are scarce. We PIT-tagged common bream Abramis brama, European perch Perca fluviatilis, pike Esox lucius and roach Rutilus rutilus, released them in their lakes of origin and recaptured them after 103-3269 days. Overall, tagged fish did not differ in condition from non-tagged fish, except for small R. rutilus that had a lower length-specific body mass in one lake in 1 year. We conclude that PIT-tagging in general has negligible long-term effects on fish condition.

Computer-aided photo-identification of a rare stingray, Megatrygon microps.

A protocol for photo-identification of individual Megatrygon microps has been defined. One hundred and four identification photographs were taken between 2005 and 2019. Spot patterns on the dorsal surface were used to identify individuals. Unique scarring on eight M. microps re-observed provided an independent confirmation of pattern stability of up to 761 days. Previous studies lacked statistical testing used to validate this photo-identification approach. I3 S photo-matching software was used to successfully match images, identifying 69 individuals. A photo-matching software facilitates an open-source platform for identifying individual M. microps, allowing for better population assessments.

Best Practice Approach for Assessment of Microchip-associated Tumors in Preclinical Safety Studies: Position of the Registry of Industrial Toxicology Animal-data (RITA).

Microchip (passive radio-frequency identification device) implantation is a common and widely employed means of animal identification in laboratory animal facilities. However, these devices have been associated with tumors of the skin and subcutis in rodents. While microchip-associated tumors are rare, they pose a challenge for accurate diagnosis and documentation in preclinical toxicity studies. Documentation of these tumors should differentiate microchip-associated lesions with spontaneously occurring or test article-induced tumors. Standardizing criteria for microchip-associated lesions will aid the diagnostic process and allow for preclinical regulatory standardization. To this end, the Registry of Industrial Toxicology Animal-data have developed clear recommendations for diagnosis and documentation of microchip-associated lesions.

[Literature review of the influences on error rates when identifying equids with transponder and hot-iron branding]. / Literaturübersicht zu den Einflussfaktoren auf die Fehlerwahrscheinlichkeiten bei der Identifikation von Equiden durch Transponder und Heißbrand.

Although equids have had to be tagged with a transponder since 2009, breeding associations in Germany disagree as to which method is best suited for identification (with or without hot iron branding). Therefore, the aim of this systematic literature review was to gain an overview of how effective identification is using transponders and hot iron branding and as to which factors influence the success of identification. Existing literature showed that equids can be identified by means of transponders with a probability of 85-100%, whereas symbol brandings could be identified correctly in 78-89%, whole number brandings in 0-87% and single figures in 37-92% of the readings, respectively. The successful reading of microchips can be further optimised by a correctly operated implantation process and thorough training of the applying persons. affect identification with a scanner. The removal of transponders for manipulation purposes is virtually impossible. Influences during the application of branding marks can hardly, if at all, be standardised, but influence the subsequent readability relevantly. Therefore, identification by means of hot branding cannot be considered sufficiently reliable. Impaired quality of identification can be reduced during reading but cannot be counteracted. Based on the existing studies it can be concluded that the transponder method is the best suited of the investigated methods for clearly identifying equids, being forgery-proof and permanent. It is not to be expected that applying hot branding in addition to microchips would optimise the probability of identification relevantly.

Rapid temperature-dependent wound closure following adipose fin clipping of Atlantic salmon Salmo salar L.

Three groups of Atlantic salmon were kept at a constant temperature of 4, 10 and 14 °C. The adipose fins were removed; six fish/group were sampled at 11 subsequent time points post-clipping. Samples were prepared for histopathological examination to study the course of re-epithelization. A score sheet was developed to assess the regeneration of epidermal and dermal cell types. Wounds were covered by a thin epidermal layer between 4 and 6 h post-clipping at 10 and 14 °C. In contrast, wound closure was completed between 6 and 12 h in fish held at a constant temperature of 4 °C. By 18 h post-clipping, superficial cells, cuboidal cells, prismatic basal cells and mucous cells were discernible in all temperature groups, rapidly progressing towards normal epidermal structure and thickness. Within the observation period, only minor regeneration was found in the dermal layers. A positive correlation between water temperature and healing rates was established for the epidermis. The rapid wound closure rate, epidermal normalization and the absence of inflammatory reaction signs suggest that adipose fin clipping under anaesthesia constitutes a minimally invasive method that may be used to mark large numbers of salmon presmolts without compromising fish welfare.

An appraisal of the Maltese national livestock database with regard to bovines.

The creation of a centralised national livestock database for the islands of Malta and Gozo is of crucial importance for the identification and traceability of bovines. It is also important for compliance with the legal obligations that followed Malta's accession to the European Union in May 2004. This paper describes how the processes of identification, registration and traceability of bovines have changed since Malta's accession. The validation and integration of data originating from different departmental sections (such as the identification and registration section), the slaughterhouse and the National Veterinary Laboratory, ensures that any discrepancies are highlighted and can be investigated. Events recorded in the database enable the compliance and eligibility of bovine producers to be cross-checked when applications for European Union benefits are made. The main drawbacks and weak points of the system include financial costs for the government department, potentially late notification of the births and deaths of newborn calves, and insufficient uptake among bovine producers of the latest technology for notification of events such as births, deaths and movement of bovines.

Electronic bolus design impacts on administration.

Electronic identification of animals has become increasingly important worldwide to improve and ensure traceability. In warm and hot climates, such as Brazil, boluses can have advantages over ear tags as the internal devices reduce the risks of ear tag losses, tissue damage, and lesions on the ear. Electronic boluses, however, are often perceived as having negative characteristics, including reported difficulties of administration in small ruminants. This paper describes the factors associated with bolus design that affect the swallowing of a bolus in sheep. Other factors that might influence bolus swallowing time have also been considered. In addition, the effect of bolus design on its performance was evaluated. A total of 56 Suffolk ewes were used to assess the ease of administration and retention of 3 types of electronic ruminal boluses (mini, 11.5 × 58.0 mm and 21.7 g; small, 14.8 × 48.5 mm and 29.5 g; standard, 19.3 × 69.8 mm and 74.4 g) during a whole productive year, including pregnancy and lamb suckling. Ewe age (5.6 ± 2.3 yr) and weight (85.07 ± 8.2 kg BW) were recorded, as well as time for bolus swallowing. The deglutition of the bolus and any resulting blockages in the esophagus were monitored by visual observations. Retention and readability of the boluses were regularly monitored for d 1, wk 1, mo 1, and every mo until 1 yr. Time for bolus swallowing differed substantially with bolus type and was greater (P < 0.05) for the standard bolus (32.8 ± 6.9 s) when compared to small and mini boluses, which did not differ (8.5 ± 2.0 vs. 9.2 ± 2.7 s; P > 0.05). The bolus o.d. and length were positively correlated with swallowing time (P < 0.01). The ewe weight was negatively correlated with swallowing time (P < 0.05). At 6 mo all electronic boluses showed 100% retention rate, and at 12 mo, bolus retention was 100%, 94.5%, and 100% for mini, small, and standard boluses, respectively (P > 0.05). At 12 mo, all boluses showed 100% readability, except for small boluses, which had a readability of 94.5%. In conclusion, bolus design affected swallowing time and bolus readability. A reduction in boluses length and o.d. needs to be carried out to provide ease of administration and for boluses to be used as an effective means of electronic identification. Therefore, this study shows that adequately designed boluses are safe and suitable for identifying adult sheep and can therefore be used in hot climates.

[Comparison of susceptibility artifacts generated by microchips with different geometry at 1.5 Tesla magnet resonance imaging. A phantom pilot study referring to the ASTM standard test method F2119-07]. / Größenvergleich von Suszeptibilitätsartefakten verursacht durch Mikrochips unterschiedlicher Geometrie am 1,5-Tesla-Magnetresonanztomographen. Phantompilotstudie unter Berücksichtigung der ASTM-Standardtestmethode F2119-07.

OBJECTIVE: Ferromagnetic material in microchips, used for animal identification, causes local signal increase, signal void or distortion (susceptibility artifact) on MR images. To measure the impact of microchip geometry on the artifact's size, an MRI phantom study was performed. MATERIAL AND METHODS: Microchips of the labels Datamars®, Euro-I.D.® and Planet-ID® (n = 15) were placed consecutively in a phantom and examined with respect to the ASTM Standard Test Method F2119-07 using spin echo (TR 500 ms, TE 20 ms), gradient echo (TR 300 ms, TE 15 ms, flip angel 30°) and otherwise constant imaging parameters (slice thickness 3 mm, field of view 250 x 250 mm, acquisition matrix 256 x 256 pixel, bandwidth 32 kHz) at 1.5 Tesla. Image acquisition was undertaken with a microchip positioned in the x- and z-direction and in each case with a phase-encoding direction in the y- and z-direction. The artifact size was determined with a) a measurement according to the test method F2119-07 using a homogeneous point operation, b) signal intensity measurement according to Matsuura et al. and c) pixel counts in the artifact according to Port and Pomper. RESULTS: There was a significant difference in artifact size between the three microchips tested (Wilcoxon p = 0.032). A two- to three-fold increase in microchip volume generated an up to 76% larger artifact, depending on the sequence type, phase-encoding direction and chip position to B0. CONCLUSION AND CLINICAL RELEVANCE: The smaller the microchip geometry, the less is the susceptibility artifact. Spin echoes (SE) generated smaller artifacts than gradient echoes (GE). In relation to the spatial measurement of the artifact, the switch in phase-encoding direction had less influence on the artifact size in GE- than in SE-sequences. However, the artifact shape and direction of SE-sequences can be changed by altering the phase. The artifact size, caused by the microchip, plays a major clinical role in the evaluation of MRI from the head, shoulder and neck regions.

Surgical wound healing in radio-tagged adult Pacific lamprey Entosphenus tridentatus held on different substrata.

Radio-tagged adult Pacific lamprey Entosphenus tridentatus held in a raceway with Plexiglas-lined walls and bottom healed more slowly and retained sutures longer than fish held in an all-concrete raceway or one with Plexiglas walls and a cobble-lined bottom. On all substrata, healing depended on when sutures were lost, and fish that lost their sutures in <14 days post-surgery healed faster than those that kept sutures longer. Long-term suture retention led to tissue trauma, infection and poor survival.

Short communication: Long-term survival of flag eartags on an Israeli dairy farm.

Two-plated self-piercing eartags were first developed in the 19th century, but information on their retention rates is scarce. A method is presented that facilitates estimation of eartag retention rate by using a random sample of cows that initially had 2 tags (1 on each ear) placed for identification and at least 1 of which survived. Striving to adopt the European Union standard for cattle ear tagging, the Israeli veterinary service conducted a field test to evaluate the performance of plastic eartags under the conditions of a typical Israeli dairy farm. The initial sample (n=900 cows) was tagged on a single farm. Retention rates were estimated based on the ratio between the observed numbers of cows with 1 or 2 eartags in the surviving group (n=97 cows). Based on this long-term (>3 yr) field test, the highest yearly retention of flag eartags (0.89±0.03) was lower than expected (0.98). Tag design and on-farm management were key factors affecting tag retention. A better design of the feedline yoke system in the feeding area, avoiding slits that can entangle the eartags, would help increase tag retention.