Infant neurodevelopment and behavior in Guadeloupe after lead exposure and Zika maternal infection during pregnancy.

BACKGROUND: Prenatal lead exposure is known to have neurotoxic effects on the developing fetus, while some viral infections may have a tropism for the central nervous system. Our objective was to study whether the effects of prenatal lead exposure on infant development and behaviors at 18 months of age are modified by the occurrence of a maternal infection to Zika virus (ZIKV) during pregnancy. METHODS: During the ZIKV epidemic in Guadeloupe in 2016 a cohort of pregnant women was set up. Blood samples (pregnancy, childbirth and cord) (n = 297) enabled us to measure blood lead levels aimed to determine prenatal lead exposure and the likelihood of maternal infection during pregnancy (ZIKV status + vs -). The 18 months "Ages and Stages Questionnaire" (ASQ) was used to generate scores for global development, fine and gross motor skills, communication, problem solving, and personal-social skills. The questions from a longitudinal cohort study conducted in Canada (Québec) were used to generate hyperactivity, opposition, inattention and physical aggression scores. Associations were tested by multivariate linear regressions. RESULTS: Prenatal lead exposure was associated with delays in neurodevelopment at 18 months, reflected by lower scores in ASQ totals, and in the fine motor and problem-solving domains. Some of these associations appeared to be sex-specific, observed almost exclusively in boys (ASQ total, fine motor and personal-social scores). Prenatal lead exposure was not associated with behavioral scores. ZIKV infection during pregnancy was associated with a lower fine motor ASQ score, and higher scores for hyperactivity, opposition and physical aggression. Significant interaction between prenatal lead exposure and ZIKV status was observed with a lower personal-social score in ZIKV (-) only, and for hyperactivity and inattention scores, though some of these interactions (ASQ personal-social score, inattention score) were no longer significant when children with microcephaly were excluded from the analyses. DISCUSSION/CONCLUSION: Our study confirms previous findings of associations between prenatal exposure to lead at low levels and adverse neurodevelopmental outcomes during infancy and the particular vulnerability of boys. It suggests associations between ZIKV infection during pregnancy and adverse effects on a number of neurodevelopmental functions (fine motor function) and behaviors (opposition, hyperactivity), that need to be confirmed at later age. There is no strong evidence of interaction between ZIKV infection and lead exposure but both prenatal risk factors may affect fine motor function.

Simultaneous exposure to both Zika virus and household insecticides during pregnancy, and fetal growth and infant developmental behavior outcomes at 18 months, in Guadeloupe.

BACKGROUND: Perinatal infection with Zika virus (ZIKV) could result in adverse growth, developmental and behavioral outcomes, while insecticides used to control mosquitoes are neurotoxic. OBJECTIVES: We aim to study the role played by exposure during pregnancy to both ZIKV and household insecticides in newborn health, development and behavior at age of 18 months. METHODS: Maternal and cord blood samples from a cohort of pregnant women (created during Guadeloupe's Zika epidemic of 2016) were used to identify ZIKV infection during pregnancy. A self-administered questionnaire at birth documented prenatal household use of insecticides. Birth weight and head circumference were collected from maternity records (n = 708). Infant development and behaviors were documented at 18 months of age through the Ages and Stages Questionnaire and the Quebec Longitudinal Study of Child Development (n = 409). Logistic and linear regression models were performed, taking into account confounding factors. RESULTS: Use of household insecticides was associated with smaller head circumference and lower birth weight among newborns from mothers not exposed to ZIKV: 0.3 cm (95% CI: 0.6, 0) and -82 g (95% CI: 165, 0), respectively. Similar decreases were observed with ZIKV exposure among mothers not reporting household insecticides use, and with presence of both exposures. The combined presence of ZIKV exposure and insecticide use was associated with lower ASQ fine motor scores (-3.9; 95% CI: 7.3, -0.4), and higher hyperactivity scores (0.8; 95% CI: 0.0, 1.5), compared to no exposure to either. A higher opposition score was observed in association with ZIKV exposure among non-users of insecticide (0.6; 95% CI: 0.0, 1.2). CONCLUSION: Adverse neurodevelopmental outcomes at 18 months of age were observed with prenatal ZIKV exposure, and with higher magnitude when mothers reported use of household insecticides. At birth, rates of adverse fetal growth were however similar for the combined presence of exposure and either of the exposures.

Prenatal exposure to legacy contaminants and visual acuity in Canadian infants: a maternal-infant research on environmental chemicals study (MIREC-ID).

BACKGROUND: Prenatal exposure to environmental contaminants can have deleterious effects on child development. While psychomotor, cognitive and behavioural outcomes have been investigated in relation to chronic exposure, the associations with visual functions remains unclear. The present study's aim was to assess the associations of prenatal exposure to legacy persistent organic pollutants and heavy metals with visual acuity in Canadian infants. The potential protective effects of selenium against mercury toxicity were also examined. METHODS: Participants (mean corrected age = 6.6 months) were part of the Maternal-Infant Research on Environmental Chemicals (MIREC) study. Concentrations of polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), lead and mercury were measured in maternal blood during pregnancy, as well as in the cord blood. The Teller acuity card test (TAC) (n = 429) and the visual evoked potentials in a sub-group (n = 63) were used to estimate behavioural and electrophysiological visual acuity, respectively. Multivariable linear regression models were used to investigate the relationship between exposure to each contaminant and visual acuity measures, while controlling for potential confounders. Breastmilk selenium, which was available for about half of the TAC and VEP samples, was also taken into account in the mercury models as exploratory analyses. RESULTS: We observed no significant associations between exposure to any contaminants and TAC. Analyses revealed a negative trend (p values < 0.1) between cord blood lead and mercury and electrophysiological visual acuity, whereas PCB and PBDE showed no association. When adding breastmilk selenium concentration to the mercury models, this association became statistically significant for cord concentrations (ß = - 3.41, 95% CI = - 5.96,-0.86), but also for blood levels at 1st and 3rd trimesters of pregnancy (ß = - 3.29, 95% CI = - 5.69,-0.88). However, further regression models suggested that this change in estimates might not be due to adjustment for selenium, but instead to a change in the study sample. CONCLUSIONS: Our results suggest that subtle, but detectable alterations of infant electrophysiological visual acuity can be identified in a population prenatally exposed to low mercury concentrations. Compared to behavioural visual acuity testing, electrophysiological assessment may more sensitive in detecting visual neurotoxicity in relation with prenatal exposure to mercury.

Androgenic and estrogenic indices in human newborns and infants: the MIREC-ID study.

Prenatal sex steroid exposure plays an important role in determining child development. Yet, measurement of prenatal hormonal exposure has been limited by the paucity of newborn/infant data and the invasiveness of fetal hormonal sampling. Here we provide descriptive data from the MIREC-ID study (n=173 girls; 162 boys) on a range of minimally invasive physical indices thought to reflect prenatal exposure to androgens [anogenital distances (AGDs); penile length/width, scrotal/vulvar pigmentation], to estrogens [vaginal maturation index (VMI) - the degree of maturation of vaginal wall cells] or to both androgens/estrogens [2nd-to-4th digit ratio (2D:4D); areolar pigmentation, triceps/sub-scapular skinfold thickness, arm circumference]. VMI was found to be associated with triceps skinfold thickness (ß=0.265, P=0.005), suggesting that this marker may be sensitive to estrogen levels produced by adipose tissue in girls. Both estrogenic and androgenic markers (VMI: ß=0.338, P=0.031; 2D:4D - right: ß=-0.207, P=0.040; left: ß=-0.276, P=0.006; AGD-fourchette - ß=0.253, P=0.036) were associated with areolar pigmentation in girls, supporting a role for the latter as an index of both androgen and estrogen exposure. We also found AGD-penis (distance from the anus to the penis) to be associated with scrotal pigmentation (ß=0.290, P=0.048), as well as right arm circumference (ß=0.462, P<0.0001), supporting the notion that these indices may be used together as markers of androgen exposure in boys. In sum, these findings support the use of several physical indices at birth to convey a more comprehensive picture of prenatal exposure to sex hormones.

Local country food sources of methylmercury, selenium and omega-3 fatty acids in Nunavik, Northern Quebec.

Country foods are central to Inuit culture and replete in selenium (Se) and long-chain omega-3 polyunsaturated fatty acids (n-3 PUFA). However, some marine country foods bioaccumulate high concentrations of methylmercury (MeHg). Se and n-3 are associated with several health benefits in Nunavik, Northern Quebec, but, recent studies show that prenatal MeHg exposure is associated with visual, cognitive and behavioral deficit later in childhood. The study objectives are to identify contemporary country food sources of MeHg, Se and long-chain n-3 PUFA in Nunavik, particularly among childbearing-age women, taking into account regional differences in consumption profiles. The contribution of different country foods to daily MeHg, Se, long-chain n-3 PUFA intake (µg/kg body weight/day) was estimated using: (i) country food consumption and blood biomarkers data from the 2004 Nunavik Health Survey (387 women, 315 men), and (ii) data on MeHg, Se, long-chain n-3 PUFA concentrations found in Nunavik wildlife species. In the region where most traditional beluga hunting takes place in Nunavik, the prevalence of at-risk blood Hg (≥ 8 µg/L) in childbearing-age women was 78.4%. While most country foods presently consumed contain low MeHg, beluga meat, not a staple of the Inuit diet, is the most important contributor to MeHg: up to two-thirds of MeHg intake in the beluga-hunting region (0.66 of MeHg intake) and to about one-third in other regions. In contrast, seal liver and beluga mattaaq - beluga skin and blubber - only mildly contributed to MeHg (between 0.06 and 0.15 of MeHg intake), depending on the region. Beluga mattaaq also highly contributed to Se intake (0.30 of Se intake). Arctic char, beluga blubber and mattaaq, and seal blubber contributed to most long-chain n-3 PUFA intake. This study highlights the importance of considering interconnections between local ecosystems and dietary habits to develop recommendations and interventions promoting country foods' benefits, while minimizing the risk of MeHg from beluga meat, especially for childbearing-age women.

Environmental contaminants and human health in the Canadian Arctic.

The third Canadian Arctic Human Health Assessment conducted under the Canadian Northern Contaminants Program (NCP), in association with the circumpolar Arctic Monitoring and Assessment Programme (AMAP), addresses concerns about possible adverse health effects in individuals exposed to environmental contaminants through a diet containing country foods. The objectives here are to: 1) provide data on changes in human contaminant concentrations and exposure among Canadian Arctic peoples; 2) identify new contaminants of concern; 3) discuss possible health effects; 4) outline risk communication about contaminants in country food; and 5) identify knowledge gaps for future contaminant research and monitoring. The nutritional and cultural benefits of country foods are substantial; however, some dietary studies suggest declines in the amount of country foods being consumed. Significant declines were found for most contaminants in maternal blood over the last 10 years within all three Arctic regions studied. Inuit continue to have the highest levels of almost all persistent organic pollutants (POPs) and metals among the ethnic groups studied. A greater proportion of people in the East exceed Health Canada's guidelines for PCBs and mercury, although the proportion of mothers exceeding these guidelines has decreased since the previous assessment. Further monitoring and research are required to assess trends and health effects of emerging contaminants. Infant development studies have shown possible subtle effects of prenatal exposure to heavy metals and some POPs on immune system function and neurodevelopment. New data suggest important beneficial effects on brain development for Inuit infants from some country food nutrients. The most successful risk communication processes balance the risks and benefits of a diet of country food through input from a variety of regional experts and the community, to incorporate the many socio-cultural and economic factors to arrive at a risk management decision that will be the most beneficial in Arctic communities.

Alteration of infant attention and activity by polychlorinated biphenyls: unravelling critical windows of susceptibility using physiologically based pharmacokinetic modeling.

Pre- and postnatal exposure to polychlorinated biphenyls (PCBs) can impair behavioural function in animal models at doses within the range at which humans are commonly exposed. Yet, epidemiologic studies conducted in the US and Europe are inconsistent with regard to the developmental effects of lactational exposure to these chemicals. This inconsistency may be due to limitations in the current methodological approaches for assessing postnatal exposure to PCBs. Our study used a physiologically based pharmacokinetic (PBPK) model to simulate blood PCB levels during specific pre- and postnatal periods and to evaluate the relation of those levels to infant behaviour. A previously validated PBPK model was used to simulate infant blood PCB-153 levels at delivery and on a month-by-month basis during the first year of life for Inuit infants enrolled in a longitudinal birth cohort. Infant behaviour was assessed using the Behaviour Rating Scales (BRS) of the Bayley Scales of Infant Development (BSID-II) at 11 months of age and video coding of inattention and activity measured during the administration of the mental development subscale of the BSID-II. The estimated pre- and postnatal PCB exposure measures predicted significant increases in inattention and activity at 11 months. Whereas inattention was related to prenatal exposure, activity level, measured by non-elicited activity, was best predicted by postnatal exposure, with the strongest association obtained for simulated PCB levels during the 4th month of life. These findings are consistent with previous reports indicating PCB-induced behavioural alteration in attention and activity level. Simulated infant toxicokinetic profiles for the first year of life revealed windows of susceptibility during which PCBs may impair infant attention and activity.

Behavioural correlates of the P3b event-related potential in school-age children.

The latency and amplitude of the P3b component of event-related potentials (ERPs) have been related to behavioural performance on several attention and memory tasks in adult populations. However, the extent to which these results apply to children is unknown. This study examined the neurobehavioral correlates of the P3b component in a longitudinal sample of school-age children from Arctic Québec. Children (N=110; mean age=11.3years) were assessed on an ERP auditory oddball paradigm and a neurobehavioral evaluation targeting several aspects of cognition, including the Stewart Extended Continuous Performance Test (E-CPT), California Verbal Learning Test (CVLT), Stroop Color-Word Interference Test, and five subtests from the Wechsler Intelligence Scale for Children-Fourth edition (WISC-IV). P3b latency was positively related to reaction time measures and negatively associated with performance on the WISC-IV Digit Span Forward subtest. Amplitude of the P3b was associated with shorter completion time on the Stroop test and better delayed recognition memory performance among children who did not use semantic strategies on the CVLT. Profile analyses revealed no difference in scalp distribution of the P3b according to performance on these tests. The results are consistent with previous studies with older participants and suggest that, despite age-related differences in waveform and scalp distribution, the P3b component relates to similar neurocognitive processes in children and adults.

The relation of environmental contaminants exposure to behavioral indicators in Inuit preschoolers in Arctic Quebec.

Although lead (Pb) exposure has been identified as an important risk factor in child behavioral development, less is known regarding the relation between child behavior and exposure to polychlorinated biphenyls (PCBs) and mercury (Hg). Inuit children are particularly exposed to these chemicals and the aim of this study was to investigate the association between prenatal and postnatal exposure to Pb, PCBs, Hg and several aspects of behavioral function in Inuit preschoolers. The sample consisted of one hundred and ten 5-year-old Inuit children from Arctic Quebec. An umbilical cord blood sample was used to document prenatal exposure to Pb, PCBs and Hg. Child blood samples were collected at age 5 and the same contaminants were measured. A modified version of the Infant Behavior Rating Scale from the Bayley Scales of Infant Development-II was used to assess child behavior through examiners' ratings. Furthermore, attention, activity and emotional outcomes were assessed through behavioral coding of video recordings taken during fine motor testing. Pb exposure during childhood was associated with examiners ratings of greater impulsivity, irritability and with coding of observed inattention. Prenatal exposure to PCB 153 correlated with the examiners ratings of increased state of unhappiness and anxiety during the testing session, which was corroborated from video coding since cord PCB 153 was related to fewer manifestations of positive affects. No association was found with Hg exposure. These data corroborated those from previous Pb cohort studies and revealed an association between prenatal PCBs exposure and emotional outcomes in preschoolers.

The relation of low-level prenatal lead exposure to behavioral indicators of attention in Inuit infants in Arctic Quebec.

The aim of this study was to investigate the association between prenatal exposure to lead (Pb) and several aspects of behavioral function during infancy through examiner ratings and behavioral coding of video recordings. The sample consisted of 169 11-month-old Inuit infants from Arctic Quebec. Umbilical cord and maternal blood samples were used to document prenatal exposure to Pb. Average blood Pb levels were 4.6 mug/dL and 5.9 mug/dL in cord and maternal samples respectively. The Behavior Rating Scales (BRS) from the Bayley Scales of Infant Development (BSID-II) were used to assess behavior. Attention was assessed through the BRS and behavioral coding of video recordings taken during the administration of the BSID-II. Whereas the examiner ratings of behaviors detected very few associations with prenatal Pb exposure, cord blood Pb concentrations were significantly related to the direct observational measures of infant attention, after adjustment for confounding variables. These data provide evidence that increasing the specificity and the precision of the behavioral assessment has considerable potential for improving our ability to detect low-to-moderate associations between neurotoxicants, such Pb and infant behavior.

The relationship between lead exposure, motor function and behaviour in Inuit preschool children.

The main focus of this study was to determine the role of behaviour in the relationship between postnatal lead exposure and motor function. The sample consisted of 110 preschoolers, of age 5, from Nunavik. Lead concentration was measured at birth and at testing time. Average lead levels were of 4.9 microg/dL (0.24 micromol/L) and 5.3 microg/dL (0.26 micromol/L) for cord and child blood, respectively. Children's balance and fine motor capacities were tested. A modified version of the IBR was used to assess behaviour. Postnatal blood lead concentrations correlated positively with both impulsivity and activity. Neither pre- nor postnatal blood lead concentration correlated with attention level. The children's scores on impulsivity (I) and activity (A) were summed to create the independent variable IA, which was tested as a potential mediator between lead exposure and two dependent variables: the coefficient of covariation in alternating hand movements and transversal sway in tandem position. Mediation was significant only for the latter variable. IA and attention were then tested as potential moderators in the relation between postnatal lead exposure and motor function. No significant interaction between independent variables could be observed. These results do not support the hypothesis that, at low levels of postnatal exposure, lead acts indirectly on motor function via behaviour. However, IA does act as a mediator in the relationship between postnatal blood lead concentration and transversal sway in tandem position.

Human health implications of environmental contaminants in Arctic Canada: A review.

The objectives of this paper are to: assess the impact of exposure to current levels of environmental contaminants in the Canadian Arctic on human health; identify the data and knowledge gaps that need to be filled by future human health research and monitoring; examine how these issues have changed since our first assessment [Van Oostdam, J., Gilman, A., Dewailly, E., Usher, P., Wheatley, B., Kuhnlein, H. et al., 1999. Human health implications of environmental contaminants in Arctic Canada: a review. Sci Total Environ 230, 1-82]. The primary exposure pathway for contaminants for various organochlorines (OCs) and toxic metals is through the traditional northern diet. Exposures tend to be higher in the eastern than the western Canadian Arctic. In recent dietary surveys among five Inuit regions, mean intakes by 20- to 40-year-old adults in Baffin, Kivalliq and Inuvialuit communities exceeded the provisional tolerable daily intakes (pTDIs) for the OCs, chlordane and toxaphene. The most recent findings in NWT and Nunavut indicate that almost half of the blood samples from Inuit mothers exceeded the level of concern value of 5 microg/L for PCBs, but none exceeded the action level of 100 microg/L. For Dene/Métis and Caucasians of the Northwest Territories exposure to OCs are mostly below this level of concern. Based on the exceedances of the pTDI and of various blood guidelines, mercury and to a lesser extent lead (from the use of lead shot in hunting game) are also concerns among Arctic peoples. The developing foetus is likely to be more sensitive to the effects of OCs and metals than adults, and is the age groups of greatest risk in the Arctic. Studies of infant development in Nunavik have linked deficits in immune function, an increase in childhood respiratory infections and birth weight to prenatal exposure to OCs. Balancing the risks and benefits of a diet of country foods is very difficult. The nutritional benefits of country food and its contribution to the total diet are substantial. Country food contributes significantly more protein, iron and zinc to the diets of consumers than southern/market foods. The increase in obesity, diabetes and cardiovascular disease has been linked to a shift away from a country food diet and a less active lifestyle. These foods are an integral component of good health among Aboriginal peoples. The social, cultural, spiritual, nutritional and economic benefits of these foods must be considered in concert with the risks of exposure to environmental contaminants through their exposure. Consequently, the contamination of country food raises problems which go far beyond the usual confines of public health and cannot be resolved simply by risk-based health advisories or food substitutions alone. All decisions should involve the community and consider many aspects of socio-cultural stability to arrive at a decision that will be the most protective and least detrimental to the communities.

Monitoring of umbilical cord blood lead levels and sources assessment among the Inuit.

Analyses completed on samples collected between 1993 and 1996 showed that about 7% of 475 Inuit newborns from northern Quebec (Canada) had a cord blood lead concentration equal to or greater than 0.48 micromol/l, an intervention level adopted by many governmental agencies. A comparison between the cord blood lead isotope ratios of Inuit and southern Quebec newborns showed that lead sources for these populations were different. Our investigation suggests that lead shots used for game hunting were an important source of lead exposure in the Inuit population. A cohort study conducted in three Inuit communities shows a significant decrease of cord blood lead concentrations after a public health intervention to reduce the use of lead shot. Lead shot ammunition can be a major and preventable source of human exposure to lead.

Prenatal exposure of the northern Québec Inuit infants to environmental contaminants.

The Inuit population residing in Nunavik (northern Québec, Canada) relies on species from the marine food web for subsistence and is therefore exposed to high doses of environmental contaminants such as polychlorinated biphenyls and methylmercury and to a lesser extent lead. In view of the neurotoxic properties of these substances following developmental exposure, we initiated a study on infant development in this remote coastal population. Here we report the magnitude of prenatal exposure to these contaminants and to selective nutrients in Inuit mothers and their newborns who were recruited on the Hudson Bay coast. We conducted interviews during the women's pregnancies and at 1 and 11 months postpartum and collected biological samples for mercury, lead, polychlorinated biphenyls (PCBs), and chlorinated pesticides analyses as well as selenium and N-3 polyunsaturated fatty acids (n3-PUFA). Cord blood, maternal blood, and maternal hair mercury concentrations averaged 18.5 microg/L, 10.4 microg/L, and 3.7 microg/g, respectively, and are similar to those found in the Faroe Islands but lower than those documented in the Seychelles Islands and New Zealand cohorts. Concentrations of PCB congener 153 averaged 86.9, 105.3, and 131.6 microg/kg (lipids) in cord plasma, maternal plasma, and maternal milk, respectively; prenatal exposure to PCBs in the Nunavik cohort is similar to that reported in the Dutch but much lower than those in other Arctic cohorts. Levels of n3-PUFA in plasma phospholipids and selenium in blood are relatively high. The relatively low correlations observed between organochlorine and methylmercury concentrations may make it easier to identify the specific developmental deficits attributable to each toxicant. Similarly, the weak correlations noted between environmental contaminants and nutrients will facilitate the documentation of possible protective effects afforded by either n3-PUFA or selenium against neurotoxic contaminants.

Determinants of polychlorinated biphenyls and methylmercury exposure in inuit women of childbearing age.

The objectives of this study were to to identify maternal characteristics associated with traditional food consumption and to examine food items associated with polychlorinated biphenyls (PCBs) and mercury body burden in pregnant Inuit women from Northern Québec. We interviewed women from three communities at mid-pregnancy and at 1 and 11 months postpartum. We measured PCBs, Hg, and selenium in maternal blood; Hg was also measured in maternal hair. The women reported eating significant amounts of fish, beluga muktuk/fat, seal meat, and seal fat. Although consumption of fish and seal was associated with lower socioeconomic status, consumption of beluga whale was uniform across strata. Fish and seal meat consumption was associated with increased Hg concentrations in hair. Traditional food intake during pregnancy was unrelated to PCB body burden, which is more a function of lifetime consumption. This study corroborated previous findings relating marine mammal and fish consumption to increased Hg and selenium body burden. Despite widespread knowledge regarding the presence of these contaminants in traditional foods, a large proportion of Inuit women increased their consumption of these foods during pregnancy, primarily because of pregnancy-related changes in food preferences and the belief that these foods are beneficial during pregnancy.

Prenatal exposure of Canadian children to polychlorinated biphenyls and mercury.

This article documents the exposure to environmental contaminants within sub-groups of the Canadian population who are considered to be at risk as a result of the food they eat. We measured the concentrations of polychlorinated biphenyls (PCBs) and mercury in the blood drawn from the umbilical cords of newborns in various Aboriginal communities, in a coastal community and in the general population. Average concentrations of Aroclor 1260 ranged between 0.3 and 2.0 micrograms/L and were clearly highest among the Inuit of Nunavik and Baffin Island and among the Montagnais of Quebec. In these groups, we found contaminant levels in the blood of newborns that exceed the threshold beyond which cognitive impairments are expected to result. Average concentrations of mercury ranged between 1.0 and 14.2 micrograms/L; the Inuit of Nunavik and the NWT exhibited the highest exposure levels. A portion of the Nunavik and NWT Inuit had concentrations beyond the critical threshold for the appearance of neurological consequences. The variations in exposure levels resulted from the different nutritional practices of these Canadian sub-groups.

On the linkages between chromophore and protein in biliproteins, VII. Amino acid sequence in the chromophore regions of C-phycoerythrin from Pseudanabaena W 1173 and Phormidium persicinum.

Bilipeptides from all chromophore regions were prepared by trypsin digestion of C-phycoerythrin from Pseudanabaena W 1173 and Phormidium persicinum. Analytical separation and quantitative determination of bilipeptides was achieved by isoelectric focusing, preparative isolation by gel chromatography and ion-exchange chromatography. Amino acid analysis revealed cysteine as the only amino acid common to all chromopeptides. Amino acid sequences were determined by Edman degradation and the dansyl-Edman technique. Sequences are different in all 5 and 6 chromophore regions, respectively. Possible homologies are discussed. A thioether linkage between ring A of the chromophore and cysteine was found in the bilipeptides (as before in biliproteins). A second linkage (serine ester) was found in only one peptide (CM 4.I from Pseudanabaena W 1173). This peptide absorbs as cation at a longer wavelength (559 nm) than the other bilipeptides (542 - 550 nm).

Chromophore content of C-phycoerythrin from various Cyanobacteria.

The molar extinction coefficient for phycoerythrobilin (l a) was calculated by two independent methods. It is different from that of the cleaved chromophore, phycobiliviolin (2). By unfolding with urea or tryptic digestion, the chromophore absorption of C-phycoerythrin (PE) was determined free of any protein influence. The chromophore content of PE from various Cyanobacteria was determined with these data to be either 5 or 6, depending on the organism. This corresponds to a chromophore distribution over phycoerythrin subunits alpha:beta of either 2:3 or 2:4. The phylogenetic significance of varying chromophore content is discussed.