<i>In situ</i> video monitoring of by-catch interactions within commercial rock lobster (<i>Jasus edwardsii</i>) fishing traps

<p>The incidental capture of non-target and undersized target species during commercial fishing operations has been identified as a major issue and ongoing challenge for fisheries management. In the Northern Zone rock lobster (<i>Jasus edwardsii</i>) fishery (NZRLF) of South Australia, escape gaps have been mandatory in all commercial fishing traps since 2002 to reduce non-target species (by-catch) catch rates and juvenile lobster mortality. This study used <i>in situ</i> video monitoring of by-catch species in traps with and without escape gaps to understand how by-catch species interact within commercial fishing traps. Twenty-two different by-catch species were observed entering commercial rock lobster traps, 20 of which were temperate reef finfish. Overall, on-board by-catch rates were significantly higher in hauled traps without escape gaps and were dominated by reef-dwelling finfish species, particularly <i>Meuschenia hippocrepis</i> and <i>Notolabrus tetricus</i>. However, <i>in situ</i> video monitoring within traps showed no difference in the maximum number of finfish by-catch (MaxN) in traps with or without escape gaps. We suggest that the difference in by-catch rates between hauled traps with and without escape gaps may be driven by species-specific behavioural interactions within traps prior to hauling. Overall, this study shows that the implementation of escape gaps in the NZRLF has considerably reduced fishery by-catch rates. Furthermore, the use of <i>in situ</i> video monitoring has provided a better understanding of by-catch behavioural interactions within traps, not previously identified in on-board sampling research, which may partly explain the mechanisms underpinning escape gap effectiveness.</p

Persistence of Hepatitis C Virus Traces after Spontaneous Resolution of Hepatitis C

<div><p>Hepatitis C virus (HCV) frequently causes chronic hepatitis, while spontaneous recovery from infection is infrequent. Persistence of HCV after self-limited (spontaneous) resolution of hepatitis C was rarely investigated. The current study aimed to assess incidence and robustness of HCV persistence after self-resolved hepatitis C in individuals with normal liver enzymes and undetectable virus by conventional tests. Applying high sensitivity HCV RNA detection approaches, we analyzed plasma and peripheral blood mononuclear cells (PBMC) from individuals with previous hepatitis C infection. Parallel plasma and PBMC from 24 such non-viraemic individuals followed for 0.3–14.4 (mean 6.4) years were examined. Additional samples from 9 of them were obtained 4.5–7.2 (mean 5.9) years later. RNA was extracted from 250 μl plasma and, if HCV negative, from ~5 ml after ultracentrifugation, and from <i>ex vivo</i> stimulated PBMC. PBMC with evidence of HCV replication from 4 individuals were treated with HCV protease inhibitor, telaprevir. HCV RNA was detected in 14/24 (58.3%) plasma and 11/23 (47.8%) PBMC obtained during the first collection. HCV RNA replicative strand was evident in 7/11 (63.6%) PBMC. Overall, 17/24 (70.8%) individuals carried HCV RNA at mean follow-up of 5.9 years. Samples collected 4.5–7.2 years later revealed HCV in 4/9 (44.4%) plasma and 5/9 (55.5%) PBMC, while 4 (80%) of these 5 PBMC demonstrated virus replicative strand. Overall, 6/9 (66.7%) individuals remained viraemic for up to 20.7 (mean 12.7) years. Telaprevir entirely eliminated HCV replication in the PBMC examined. In conclusion, our results indicate that HCV can persist long after spontaneous resolution of hepatitis C at levels undetectable by current testing. An apparently effective host immune response curtailing hepatitis appears insufficient to completely eliminate the virus. The long-term morbidity of asymptomatic HCV carriage should be examined even in individuals who achieve undetectable HCV by standard testing and their need for treatment should be assessed.</p></div

Total Numbers of Plasma and PBMC Samples from individuals with Spontaneous Resolution of Hepatitis C And Results on HCV Detection.

<p>PBMC, peripheral blood mononuclear cells; HCV RNA+, HCV RNA positive strand; HCV RNA-, HCV RNA negative strand; PHA, phytohemagglutinin; C5, immune cell stimulating cocktail (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0140312#sec002" target="_blank">Materials and Methods</a>); NA, not applicable.</p><p>*Tested only when the sample found HCV RNA positive strand reactive.</p><p>Total Numbers of Plasma and PBMC Samples from individuals with Spontaneous Resolution of Hepatitis C And Results on HCV Detection.</p

Expression of HCV RNA positive and negative (replicative) strands in PBMC samples obtained at two collections approximately 5 years apart from individuals with self-resolved hepatitis C.

<p>(A) HCV RNA positive strand identification using 2 μg of total RNA extracted from PBMC treated <i>ex vivo</i> with PHA or C5 (*). (B) Detection of HCV RNA negative strand in HCV RNA positive strand reactive PBMC shown in A. Samples were obtained during the first (1) or the second (2) collection at follow-up time (years) indicated under panel A. As positive controls for HCV RNA positive strand detection, serial 10-fold dilutions of rHCV UTR-E2 carrying indicated copy numbers/reaction were used (panel A). For HCV RNA negative strand detection, synthetic HCV RNA positive strand (sHCV RNA pos) and HCV synthetic RNA negative strand (sHCV RNA neg) at 10⁴ copies/reaction were used as positive and specificity controls, respectively (panel B). Water amplified in direct (DW) and nested (NW) reactions and a mock (M) extraction served as contamination controls. Positive signals demonstrated the expected 244-bp oligonucleotide fragments. Numbers under panels marked as relative DU represent relative densitometric units (DU) given by hybridization signals.</p

Clinical Characteristics and Time of Sample Collection from Individuals with Spontaneous Resolution of Hepatitis C.

<p>ALT, alanine aminotransferase; PT, prothrombin time; kPA, kilopescals; M, male; F, female; POS, positive; NEG, negative; NA, not available; NT, not tested; IDU, Intravenous drug user; Blood tx, blood transfusion,</p><p>*Assay sensitivity 20 IU/m.</p><p>^†Only plasma obtained during the first collection examined</p><p>^‡Patient with membranous glomerulonephritis.</p><p>^§Assessed by clinical test.</p><p>Clinical Characteristics and Time of Sample Collection from Individuals with Spontaneous Resolution of Hepatitis C.</p

Detection of HCV RNA in plasma obtained at two separate collections from individuals with a past episode of spontaneously resolved hepatitis C.

<p>Total RNA extracted from 250 μl or a pellet recovered after ultracentrifugation of ~5 ml plasma (*) acquired during the first (1) and the second (2) sample collections were amplified with 5'-UTR specific primers and amplicon specificity verified by NAH. Serial 10-fold dilutions of recombinant HCV 5'-UTR-E2 (rHCV UTR-E2) fragment carrying indicated copy numbers/reaction were used as positive and specificity controls. Water amplified in direct (DW) and nested (NW) reactions and a mock (M) extraction served as contamination controls. Positive signals showed the expected 244-bp oligonucleotide fragments. Numbers under the panels represent follow-up time in years after resolution of hepatitis C (upper line) and relative densitometric units (DU) given by hybridization signals (lower line).</p

Single-Nucleotide Polymorphisms in the HCV 5’-UTR Sequence from Plasma and PBMC Obtained at Two Collections Done More than 5 Years Apart from Patients after Spontaneous Resolution of Hepatitis C.

<p>The results are presented as numbers of clones in which a given mutation was identified per total number of clones tested. The designation of the nucleotide position based on the prototype 3a and 1b subgenotype sequences D1773 and D11168, respectively, from GenBank. ND, not detected; NA, not available</p><p>Single-Nucleotide Polymorphisms in the HCV 5’-UTR Sequence from Plasma and PBMC Obtained at Two Collections Done More than 5 Years Apart from Patients after Spontaneous Resolution of Hepatitis C.</p

Single-nucleotide polymorphisms in the HCV E1/E2 330-bp fragment derived from PBMC of two individuals with long-term follow-up after spontaneously resolved hepatitis C.

<p>(A) Sequence identified in PBMC after 7.4 years of observation of 15-46/F carrying HCV genotype 3a. (B) Sequence from PBMC of 20-49/F followed for 20.6 years carrying HCV genotype 1b. Seven or eight randomly selected clones were sequenced bi-directionally and their sequences compared to respective subgenotype D17763 and D11168 sequences from GenBank. The boundaries of the hypervariable region 1 (HVR1) are marked by a line. Numbers and the beginning and at the end of each reference sequence, marked as Ref, indicate nucleotide positions. Nucleotides identical to those in the reference sequence are shown as dots, different as letters, and deleted as colons.</p

Detection of HCV in Two Sequential Paired Plasma and PBMC.

<p>PBMC, peripheral blood mononuclear cells; HCV RNA+, HCV RNA positive strand; HCV RNA-, HCV RNA negative strand; PHA, phytohemagglutinin; C5, immune cell stimulating cocktail (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0140312#sec002" target="_blank">Materials and Methods</a>); POS, positive; NEG, negative; NT, not tested; NA, not available.</p><p>Detection of HCV in Two Sequential Paired Plasma and PBMC.</p

Inhibition of HCV infection in PBMC expressing initially both positive and negative strand of HCV RNA from individuals with a past spontaneous resolution of hepatitis C.

<p>PBMC naturally infected with HCV derived from asymptomatic persons followed for 7.4 (14-47/F), 16.1 (15-46/F) or 20.6 (20-49/F) years were treated (T) with 4 μM of TLV or left untreated (UT) in culture for 72 h. Except 20-49/F PBMC, the experiment was performed in duplicate. (A) HCV RNA positive strand was detected by RT-PCR with 5’-UTR-specific primers and amplicon specificity verified by NAH. (B) Virus negative (replicative) strand was identified by the strand-specific RT-PCR/NAH in which synthetic HCV RNA positive (pos) and negative (neg) strands at 10⁴ copies/reaction were used as specificity controls. Other controls were as described in the legend to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0140312#pone.0140312.g002" target="_blank">Fig 2</a>.</p