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Eduardo Rodrigues Thomaz 1*, Jayme Augusto Peres 1, Laís Cristine Werner 1, Adriano Torres
Carrasco de Oliveira 1, Meire Christina Seki 1, Fatima Maria Caetano Caldeira1, Alaor Aparecido
Almeida2 , André Luiz Filadelpho3
1 Departamento de Medicina Veterinária, Universidade Estadual do Centro-Oeste
(UNICENTRO – Campus Cedeteg), Guarapuava – PR, Brasil.
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2 CEATOX,
Evaluation of heavy metals associate to congenital malformation (Anidiano) in fetal bovine
and histological description highlighting the presence of tissue cysts of Toxoplasma gondii.
Instituto de Biociências de Botucatu – IBB, Botucatu-SP, Brasil.
Instituto de Biociências, Universidade Estadual Paulista Julio de Mesquita Filho (UNESP –
Campus Botucatu), Botucatu – SP, Brasil.
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*Author for correspondence: [email protected]. Fone: 55 42 3629 8228.
Rua Simeão Camargo Varela de Sá, 03. CEP 85040-080. Guarapuava –PR.
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ABSTRACT
Carried out a description of teratogenesis, anidiano, held in a bovine of Holstein, where an adult
female in her third pregnancy, which one was twin, generated a regular calf, without any
malformation where the animal continued his development normally and along with this was
expelled a nodular formation that had black and white coat and a structure similar to umbilical
cord to the center. The appearance of these cases are sporadic and have multifactorial origin,
wherein the histopathology exam from the anidiano, it was noted a cyst presence wrapped for
granulation tissue and the examination diagnosis was confirmed occurrence of Toxoplasma
gondii by the serological examination of the mother, also can be associated with the presence of
heavy metals who were found in concentrations of 0,080 µg/mL for cadmium and 0,220 µg/mL
for lead in fetal tissue, while in the structure such an umbilical cord were found values of 0,146
µg/mL for cadmium and levels of lead found themselves below the values can be detected by
technology from atomic absorption spectrophotometry. Thus, the presence of heavy metals, lead
and cadmium, may have favored the installation of the parasite, as these metals tend to decrease
fetal immunity.
Keywords: anidiano, bovine, heavy metals, histopathology, Toxoplasma gondii
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INTRODUTION
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It is understood by congenital anomaly every type of malformation, can it be structural or
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functional that occur during fetal development, which are associated with prenatal factors affecting
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the systems that make up all, or part of an organism (8). The same authors also write that these
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structural and functional abnormalities may occur in tissues, organs and / or systems.
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Such anomalies may have causes related to the environment as well as the nutritional state
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of the animal associated with vitamin deficiency, ingestion of toxic plants, and the incidence of
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viral agents, such as Bovine Viral Diarrhea (BVD), which can occur in the form of surge when the
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most common and present themselves sporadically, vertical diseases transmitted from female to the
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fetus, such as maternal infection by protozoa such as Toxoplasma gondii and/or Neospora caninum
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(2). The occurrence of mechanical trauma sustained during pregnancy, as also conditions of
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intrauterine life associated with the amount of amniotic fluid, the presence of teratogenic agents
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such as radiation or the use of drugs in the first third of gestation, as is the case of benzimidazoles.
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These anomalies can also be caused by genetic factors when transmitted by autosomal recessive
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genes (5,6).
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Teratogenesis are also associated with heavy metal poisoning, which are chemical
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elements in nature associated with geomorphological behavior of the earth's crust, as in case of
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lead (Pb) and cadmium (Cd) in volcanic activities, but their cumulative power for long periods
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interferes with the local fauna and flora, besides the issue of anthropogenic responsibility
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interfering through industrial and agricultural activities (4, 9). Was observed the occurrence of
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teratogenesis associated with lead, when analyze tissue samples from a inidiomos fetus bovine,
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especially in nervous tissue (7).
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CASE DESCRIPTION
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In the southwestern region of Paraná, a bovine female Holstein with five-year-old went into
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labor, eight days after the deadline, this was her third pregnancy, and in the previous two calves
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were born free of any anomaly, the parturition was properly assisted and aided, characterized as a
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simple and easily.
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The pregnancy was twins, one calf that was born continued to develop without any
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complication, but then, was expelled a nodular formation, the size of a human fist, set in
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vascularized gelatinous tissue featuring a congenital anomaly. The cow placenta showed a retention
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frame, but it was treated immediately following production normally.
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The anomalous fetus was sent to the Pathological Anatomy Laboratory in the State
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University Midwest - UNICENTRO, which was fixed in 10% buffered formalin solution for
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further dissection being described as a compact mass of firm consistency, nodular, having bristle
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and skin of two colors (black and white) (Figure 1). In the center there was a formation similar to
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the umbilical cord adhering on its opposite end with a diffuse area of fibrous and gelatinous
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tissue, cutting, was observed compact mass of dark staining with central area displaying
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papiliform formation of brownish ending at the center of the nodule (Figure 2).
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Samples from the anomalous fetus and the cord similar formation were collected to
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toxicological testing which were weighed eight grams stored into labeled plastic bags and cooled to
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-4˚C and then subjected to organic digestion with 65% nitric acid (HNO3), which was held in
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exhaustion camera and then sent for toxicological analysis by the technique of atomic absorption
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spectrophotometry in Toxicology Laboratory - CEATOX State University Paulista Julio de
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Mesquita Filho, in Botucatu campus.
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Digestion aims to eliminate the organic matter, being samples layered in nitric acid solution
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(65% HNO3) in the amount of 10 mL maintained for 24 hours, which were subsequently added 10
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mL of hydrogen peroxide (H2O2) to digestion residual fat present in the samples. The solutions
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remained for 24 hours in the refrigerator and after sent for reading Atomic Absorption
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Spectrophotometer - EAA (GBC AA932) with detectable limit (LD) less than or equal to 0.05 mcg
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(3).
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For the histology technique gathered up a fragment of the malformed fetus, and was fixed in
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10% buffered formalin solution and stained with hematoxylin and eosin for further analysis in light
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microscopy.
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DISCUSSION
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Structural defects occur due to the existence of failures during organogenesis, the
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deformations are characterized by changes in the form of an organic structure, after differentiation
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in the embryo (6). Statistics dates show that there is an overall prevalence rate of malformations
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between 0.5% and 3% for cattle, the same author also states that the incidence of congenital defects
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is greater in pure breeding, because the crossing of carrier animals is greater, aimed the purity of the
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breed (1).
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According to the classification of monsters Izidoro Geoffrey Saint Hilaire (13) the fetus is
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classified as a anidiano by having irregularly globular or ovoid shapes, covered with bristle and
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skin, maded of connective tissue, fat and branches of the umbilical cord vessels, such features could
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be observed in animals reported in this case.
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The homozygous twinning is susceptible to failures and can then generate abnormalities.
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They are described numerous degrees of union of twins, among them the fetus with two heads is
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the most commonplace. However, cases in which the spine is separated may occur, and
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occasionally born amorphous twins that are directly related to the double monsters and identical
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twins, are located attached to fetal wraps of other calves that may have an outer layer of skin
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involving adipose tissue (1). Because of this malformation be derived from a twin pregnancy and
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a calf was born without changes as described above, we can associate to that described by (12),
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where usually only one of the twins is monster, this is usually smaller in size having malformed
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or missing internal organs and for not having heart, its circulation is ensured by twin who is
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called autosita.
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With the aid of optical microscopy it was possible describe the collected sample to be made,
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in center, by connective tissue with rectilinear projections, central areas of vascularization,
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formation of papillary projections of fusiform epithelial cells and irregularly arranged round, with
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the presence of unsightly tangle of fibroblast projections, areas of dense stroma and focal
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hyperemia. While the sample edges has been described as containing diffuse connective tissue of
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uneven distribution forming a capsule, presence of keratinized epithelial cells in small quantities
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with diffuse formation of necrotic clusters as well as irregularly arranged polyploid formation. Were
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also found in optical tissue microscopy a suggestive cyst of toxoplasmosis (Figure 3), suspicion that
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was confirmed by serological examination of the matrix indicating antibody titration suggesting
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infection per Toxoplasma gondii. Parasitic diseases such as toxoplasmosis can also result in
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reproductive problems in several animal species such as cattle, goats, pigs, sheep, chicken, duck,
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peru, rabbit and so on (10). Among these problems we mention abortions, stillbirths, mummified
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fetuses and heat repetition, where the clinical signs change according to the immunity of each
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animal.
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Heavy metal analysis resulted in 0.080 µg/mL for cadmium and 0.220 µg/mL for lead in
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fetal tissue, while in the structure similar to the umbilical cord found values of 0.146 µg/mL for
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cadmium and lead levels have been found below the values that can be detected by the technique of
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atomic absorption spectrophotometry. Were found mean values of 0.12 ppm and 1.40 ppm for
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cadmium and lead, respectively, after analyze different tissues of aborted fetal calf in the seventh
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month of pregnancy (11), values that differ from those found in this case, but they are indicative of
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possible worsening of Toxoplasma gondii infection.
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Based on histology and titer of maternal antibodies can be concluded that one of the possible
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triggering factors of malformation is the parasitism by Toxoplasma gondii, but the presence of the
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heavy metals lead and cadmium can promote the installation of the parasite, once these metals,
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associated with the process, reduce fetal immunity, still being in this field of fetal toxicity of heavy
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metals there is more to be investigated.
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AKNOWLEDGMENTS
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Thanks to CEATOX for the partnership in the analysis.
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Figure 1: External view of the anomalous fetus (nodule with bristle) attached to placenta
Figure 2: About the anomalus fetus note the difference between stroma and parenchyma coloring, formation similar to
the umbilical cord, compact mass characteristics of dark staining and internally papiliform region, between stroma and
parenchyma, ending at the center.
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Figure 3: Presence os the cyst in tissue increase 10X (A) and 40X (B). Encapsulated cyst with presence of compact
mass. Amorphous tissue with presence of hyaline degeneration with fusiforms cell (fibroblasts) (#) and vacuolated
areas (*) with protein exudation
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