Zoo animals as sentinels for Schmallenberg virus monitoring in Spain.

Schmallenberg virus (SBV) is a newly emerged vector-borne pathogen that affects many domestic and wild animal species. A serosurvey was carried out to assess SBV exposure in zoo animals in Spain and to determine the dynamics of seropositivity in longitudinally sampled individuals. Between 2002 and 2019, sera from 278 animals belonging to 73 different species were collected from five zoos (A-E). Thirty-one of these animals were longitudinally sampled at three of these zoo parks during the study period. Seropositivity was detected in 28 (10.1 %) of 278 animals analyzed by blocking ELISA. Specific anti-SBV antibodies were confirmed in 20 (7.2 %; 95 %CI: 4.2-10.3) animals of six different species using virus neutralization test (VNT). The multiple logistic regression model showed that "order" (Artiodactyla) and "zoo provenance" (zoo B; southern Spain) were risk factors potentially associated with SBV exposure. Two (8.7 %) of the 31 longitudinally-sampled individuals showed specific antibodies against SBV at all samplings whereas seroconversion was detected in one mouflon (Ovis aries musimon) and one Asian elephant (Elephas maximus) in 2016 and 2019, respectively. To the best of the author's knowledge, this is the first surveillance conducted on SBV in zoos in Spain. The results confirm SBV exposure in zoo animals in this country and indicate circulation of the virus before the first Schmallenberg disease outbreak was reported in Spain. Surveillance in zoological parks could be a complementary approach to monitoring SBV activity. Further studies are warranted to assess the impact of this virus on the health status of susceptible zoo animals.

Evidence of exposure to zoonotic flaviviruses in zoo mammals in Spain and their potential role as sentinel species.

A serosurvey was carried out to assess emerging flavivirus exposure in zoo mammals in Spain and to determine the dynamics of seropositivity in species that were longitudinally sampled during the study period. Sera from 570 zoo animals belonging to 120 mammal species were collected at ten zoos (A-J) in Spain between 2002 and 2019. Twenty-one of these animals, belonging to ten different species, were sampled longitudinally at four of the zoos during the study period. Antigenically-related flavivirus antibodies were detected in 19 (3.3 %; 95 %CI: 2.0-5.2) of the 570 animals analyzed using bELISA. Seropositivity was observed in ten (8.3 %) of the 120 species tested. Five (23.8 %) of the 21 animals sampled more than once presented seropositivity in all samplings whereas seroconversion was only observed in one white rhinoceros (Ceratotherium simum). Flavivirus antibodies were found at six of the ten sampled zoos and in consecutive years between 2008 and 2018. Virus neutralization tests confirmed West Nile virus (WNV), Usutu virus (USUV) and tick-borne encephalitis virus (TBEV) infection in ten (1.8 %; 95 %CI: 0.7-2.8), five (0.9 %; 95 %CI: 0.1-1.6) and one (0.2 %; 95 %CI: 0.0-0.5) animal, respectively. Antibodies against Meaban virus (0 %; 95 %CI: 0.0-0.7 %) were not found in the tested sera. The results demonstrate WNV, USUV and TBEV exposure in zoo mammals, which may be of public health and conservation concern. Seropositivity to WNV and USUV was detected in regions where these viruses have not been reported previously. Anti-WNV antibodies found in zoo animals sampled in 2009 point to WNV circulation at least one year before the first outbreaks were reported in horses and humans in Spain. Our results indicate that zoo mammals could be useful sentinel species for monitoring emerging flavivirus activity in urban areas.

Monitoring of the novel rabbit haemorrhagic disease virus type 2 (GI.2) epidemic in European wild rabbits (Oryctolagus cuniculus) in southern Spain, 2013-2017.

Rabbit hemorrhagic disease (RHD) is a highly infectious disease in European rabbits (Oryctolagus cuniculus), caused by a virus belonging to the genus Lagovirus (RHDV; family Caliciviridae). In 2010, a new genotype of RHDV (RHDV2 or RHDVb, currently designated GI.2) emerged in France, affecting both domestic rabbits, even those vaccinated for the classical RHDV genotypes (currently designated GI.1) and wild rabbits. GI.2 was subsequently identified in other European countries. The aim of the present study was to monitor the GI.2 epidemic in wild rabbits in Andalusia (southern Spain) during the period 2013-2017. At the beginning of summer 2013, high mortalities were detected in wild rabbit populations in southern Spain. A total of 96 affected hunting or protected areas were surveyed. The first outbreak was observed on June 2013. The number of outbreaks sharply increased in 2013 and 2014, with a decreasing trend being observed during the following years. The spatial distribution of GI.2 was not homogeneous, since most of the detected outbreaks were concentrated in the western part of Andalusia. The outbreaks peaked in winter and spring and have been detected in the last five consecutive years, which suggests endemic circulation of GI.2 in wild rabbit populations in Spain. A total of 190 dead rabbits from 87 of the 96 areas surveyed were collected during the study period. Mortality affected rabbits of different age classes, including kittens. RT-PCR confirmed the presence of GI.2 RNA in the livers of 185 of the 190 (97.4%) rabbits. Phylogenetic analysis performed on eleven samples collected in different provinces of Andalusia between 2013 and 2017, showed high nucleotide identity with GI.2 strains Spain, France and Portugal. The results constitute an important step in understanding of the emergence and spread of GI.2 in this country and will provide valuable information for the development of surveillance programs in Europe.

Malignant colonic adenomas. Therapeutic criteria. Long-term results of therapy in a series of 42 patients in our healthcare area

Objective: a) to evaluate the appropriateness of histological criteria as proposed by Morson as indicators for surgery; and b) to compare the adequacy of Morson’s criteria with Haggitt’s levels as indicators for surgery in the case of malignant sessile lesions. Material and methods: we carried out a prospective, protocolized study of 42 patients with polyps with invasive carcinoma (IC) who underwent colonoscopic polypectomy from 1979 through 2008. We applied the histological criteria proposed by Morson to all the patients included in the series. Results: we treated 24 polyps with IC and favorable histological criteria (FC) and 18 polyps showing unfavorable histological criteria (UC). All polyps with FC were treated by means of colonoscopic polypectomy. None of the patients showed signs of disease after a mean follow-up period o f 9.67 yrs. Patients with polyps with UC were recommended to undergo surgery. The presence of unfavorable histological criteria in the polypectomy sample was clearly associated with an unfavorable patient outcome and showed a 100% sensitivity level, a negative prognostic value (NPV) of 100%, and a false negative (FN) percentage of 0% for the prognosis of the disease. We found 12 sessile polyps (Haggitt’s level 4). Colonoscopic polypectomy was the treatment employed in 9 out of 12 cases. All patients are free from disease (mean follow- up 7.3 yrs). If Haggitt’s level criteria had been applied, all 12 patients would have undergone surgery. This means 58% more patients than following Morson’s criteria. Conclusions: Morson’s criteria are considered an adequate diagnostic tool for the indication of surgery in patients with malignant adenomas. Haggitt’s invasion levels do not accurately discriminate the necessity for surgery in case of malignant sessile lesions(AU)

Malignant colonic adenomas. Therapeutic criteria. Long-term results of therapy in a series of 42 patients in our healthcare area.

OBJECTIVE: a) to evaluate the appropriateness of histological criteria as proposed by Morson as indicators for surgery; and b) to compare the adequacy of Morson s criteria with Haggitt s levels as indicators for surgery in the case of malignant sessile lesions. MATERIAL AND METHODS: we carried out a prospective, protocolized study of 42 patients with polyps with invasive carcinoma (IC) who underwent colonoscopic polypectomy from 1979 through 2008. We applied the histological criteria proposed by Morson to all the patients included in the series. RESULTS: we treated 24 polyps with IC and favorable histological criteria (FC) and 18 polyps showing unfavorable histological criteria (UC). All polyps with FC were treated by means of colonoscopic polypectomy. None of the patients showed signs of disease after a mean follow-up period o f 9.67 yrs. Patients with polyps with UC were recommended to undergo surgery. The presence of unfavorable histological criteria in the polypectomy sample was clearly associated with an unfavorable patient outcome and showed a 100% sensitivity level, a negative prognostic value (NPV) of 100%, and a false negative (FN) percentage of 0% for the prognosis of the disease. We found 12 sessile polyps (Haggitt s level 4). Colonoscopic polypectomy was the treatment employed in 9 out of 12 cases. All patients are free from disease (mean follow-up 7.3 yrs). If Haggitt s level criteria had been applied, all 12 patients would have undergone surgery. This means 58% more patients than following Morson s criteria. CONCLUSIONS: Morson s criteria are considered an adequate diagnostic tool for the indication of surgery in patients with malignant adenomas. Haggitt s invasion levels do not accurately discriminate the necessity for surgery in case of malignant sessile lesions.

[Recurrent acute pancreatitis caused by ampullar intestinal duplication filled with calculi]. / Pancreatitis aguda recurrente por duplicación intestinal ampular repleta de cálculos.

OBJECTIVES: Anatomical anomalies are recognised but rare causes of acute pancreatitis. We report a case or a 33-year-old woman with recurrent attacks of acute pancreatitis associated with a ampullar intestinal duplication. MATERIAL AND METHOD: This patient was clinically studied, and a barium contrast study of the duodenum showed a filling defect in the second part of the duodenum, that was thought to be a periampullary lipoma. Endoscopic retrograde cholangiopancreatography was scheduled, but cannulation of the papillary orifice failed. RESULTS: Laparotomy and surgical resection of the lesion were performed. Pathological study of the specimen revealed a duodenal duplication filled with multiple calculi. CONCLUSION: This entity should be considered among the lesions that can cause recurrent acute pancreatitis.

Passage pressure across the uterotubal junction during the 4 phases of the oestrous cycle in the rat.

The closure strength of the uterotubal junction (UTJ) was measured in 90 2.5 month old cycling female Wistar rats, assigned to 9 groups of 10 each. The passage pressure in both left and right UTJ were registered in pro-oestrus, oestrus, metoestrus and dioestrus phases. Five other groups were formed with rats which had been treated for 3 weeks with oestradiol (group E) or medroxiprogesterone acetate (group P), and with animals which had been ovariectomized 75 days prior to registering the pressure (ovariectomized group) or ovariectomized and treated with oestradiol (group Ov-E) or medroxiprogesterone acetate (group Ov-P). The passage pressure of the right and left UTJ in each animal was similar in each of the groups. The pressures obtained in the oestrus phase or in the E and Ov-E groups were significantly smaller than those registered in dioestrus or in the P and Ov-P groups, and only slightly smaller than that obtained in the pro-oestrus and metoestrus phases. The results indicate that the swelling of the mucosa mediated by oestrogens is not the cause of the tubal locking in the rat.

Anastomosis of uterine serosa with cyanoacrylate versus suture in rats.

Experimental end-to-end uterine horn anastomosis was performed in 20 Wistar rats with the conventional technique of microsurgery, with n-butyl-cyanoacrylate and with cyanoacrylate with the addition of one or two microsutures. Only in the case of conventional anastomosis and of cyanoacrylate in combination with two microsutures, was the patency rate 100%, whereas in the case of uterine horns anastomosed with cyanoacrylate alone, the patency rate was 80%. Histologic findings revealed the presence of tissue adhesive encysted in the serosa with a moderate inflammatory reaction around it, remaining two months after surgery. On the 60th day, the muscularis layer was only continuous in the conventional and cyanoacrylate with two microsutures groups. Our findings indicate that tissue adhesive and two sutures used together may offer a viable alternative to sutures for genital duct anastomosis.

Study of two simplified microsurgical techniques for uterine horn anastomosis in rat.

This study compares two simplified techniques, which use only 2 and 4 sutures respectively, with a conventional technique with 8 sutures for fallopian tube anastomosis. Experimentally these techniques were performed on the uterine horns of fifteen female rats. A 100% patency rate was obtained with all three techniques. No difference in the mucosal, muscular and serosal regeneration was observed in the three groups at 10, 20 and 60 days. Two months after surgery, the serosa, muscularis and mucosal layers were completely continuous in all groups. The operating with 2 sutures (5'30" +/- 1'10") was significantly less than with 4 (9'09" +/- 0'55", P < 0.05 ANOVA) and 8 sutures (15'12" +/- 1'41", P < 0.05 ANOVA). A minimum inflammatory reaction to sutures was observed in all three groups at 60 days after surgery. The results suggest that with 2 sutures are all that is needed for tissue repair.

Postoperative adhesion after uterine horn surgery in the rat, with the use of TC7.

The influence of TC7 in adhesion prevention was tested on 24, 2.5-month-old Wistar rats. Their right and left uterine horns were homogeneously allotted into four experimental groups, each of 12 horns. The groups were as follows: Group C (n = 12), that underwent cuts along 1 cm in the serosal layer; Group TC (n = 12), where the uterine horn was only surrounded with 3 layers of TC7, 1 x 1 cm in size; Group C+TC (n = 12) in which the serosal cuts were covered with TC7, as in group TC and an untouched control group (n = 12). All uterine horns were kept in uniform contact with the intestinal serosa layer by means of two 8/0 nylon stitches spaced 2 cm apart, so that the adhesion surfaces were the same in all groups. Lengths in mm and severity adhesions (0 = no adhesion to 3 = severe adhesion) were recorded. The injury in group C caused extensive (8.83 +/- 4.77 mm) and dense adhesions (score 2.42 +/- 1.11) in 83.33% of the cases. No significant differences were observed compared with the C+TC group (8.08 +/- 5.45 mm, score 2.25 +/- 1.30; P > 0.05, ANOVA). Significantly, more adhesions were observed on the C and C+TC Groups than in the control (1.00 +/- 2.27 mm, score 0.16 +/- 0.37) and TC (3.08 +/- 3.38 mm, score 1.00 +/- 1.15) groups (P < 0.05, ANOVA). TC7 does not appear to be an effective agent for preventing postoperative adhesions in this animal model.

Uterine horn replacement in the rat using denervated and devascularized jejunum.

In the hope of identifying suitable material for reconstructive tubal and uterine surgery, the authors have replaced a portion of the uterine horns of two groups of Wistar rats (n = 10) with normal (VAS group) or with denervated and devascularized jejunal grafts (NVAS group). All grafted horns stayed patent. No pregnancy was observed in the uterine horns operated on. The light microscopic appearance of the grafts was studied and compared by ANOVA. Three months after surgery in the VAS group, the number (51.4 +/- 9.5) and height (309.8 +/- 44.7 microns, P less than 0.05) of the villus, the Lieberkühn cripts height (156.9 +/- 34.9 microns) and the number of goblet cells per cript (7.2 +/- 1.2, P less than 0.05) had decreased in relation to their corresponding control jejunal fragments (57.7 +/- 5.4; 345.1 +/- 18.0 microns; 192.2 +/- 40.8 micron; 12.4 +/- 1.9, respectively). Three months after surgery, the intestinal structure was maintained in the anastomosis areas of the NVAS group, therefore the mucus production of the jejunal-graft mucosa was not completely eliminated. No villus or crypts were observed, but a monostratified cubic-cell-type epithelium at the NVAS jejunal graft mid-portion, along 58.1 +/- 12.9% of the total length of the intestinal graft. Under this epithelium a connective tissue, like a non-glandular submucosa, was apparent.

Structural changes of a devascularizated colonic graft when its replaces the rat uterine horn.

A one centimeter length of the left uterine horn in two groups of Wistar rats (n = 10) was replaced with normal (VAS group) or with denervated and devascularizated (NVAS group) colonic grafts. All animals maintained pregnancy in the right control horns but not in the grafted horns. At 40, 60 and 90 days after surgery, the light microscopic appearance of the autografts was studied. In the VAS group, and with respect to the last period, the number (5.5 +/- 0.7) and height (1.0 +/- 0.1 mm, P < 0.05 ANOVA) of the folds, the intestinal glands height (160.2 +/- 21.2 mu, P < 0.05 ANOVA) and the number of globet cells per gland (26.6 +/- 4.2, P < 0.05 ANOVA) had decreased in relation to the colon control (6.0 +/- 0.7, 1.4 +/- 0.1, 251.7 +/- 31.8, 42.6 +/- 5.2 respectively). A similar intestinal structure to that described above was observed in the anastomosis areas of the NVAS group, and therefore a decreased mucus production was maintained in this areas. No folds or intestinal glands were observed, but a monoestratified cubic epithelial cells type was observed along 63.7 +/- 4.1% of the NVAS colonic graft center. Under this epithelium a connective tissue, like a non-glandular submucosa, was obtained.